THE RESPONSE OF INBRED LINES OF CHICKENS TO LYMHOID TTIMOR TRANSPLANTS ByAhmad H * E l D a r d ir y A THESIS S u b m itted t o t h e S c h o o l o f G raduate S t u d ie s o f M ich igan S t a t e C o lle g e o f A g r ic u lt u r e and A p p lie d S c ie n c e i n p a r t i a l f u l f i l l m e n t o f t h e r e q u ir e m e n ts fo r th e degree o f DOCTOR OF miLOSOPHY D epartm ent o f A nim al H usbandry 1950 ProQ uest Number: 10008288 All rights reserved INFO RM ATION TO ALL USERS The quality o f this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a com plete m anuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. uest ProQ uest 10008288 Published by ProQuest LLC (2016). Copyright of the Dissertation is held by the Author. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code M icroform Edition © ProQ uest LLC. ProQ uest LLC. 789 East Eisenhow er Parkway P.O. Box 1346 Ann Arbor, Ml 4 8 1 0 6 - 1346 THE RESPONSE OF INBRED LINES OF CHICKENS TO LYB5PHOID TTJMOR TRANSPLANTS 3 3 4 4 9 3 TABLE OF CONTENTS In tr o d u c tio n : L it e r a t u r e c it e d * 1. D e fin itio n . 2. I n h e r it a n c e i n r e l a t i o n t o d i s e a s e . 3. I n h e r it a n c e i n r e l a t i o n t o tu m ors i n m ic e . 4. N a t u r a lly o c c u r r in g lym p h om atosis* 5. E x p e r im e n ta lly t r a n s p la n t e d ly m p h o m a to sis. M a t e r ia ls and M ethods 1* S o u rce o f m a t e r ia l f o r in o c u lu m . 2. P r e p a r a tio n o f in o c u lu m . 3. A d m in is tr a tio n o f in o c u lu m . 4. R e c ip ie n ts . 5. C o lle c tio n o f d a ta . 6. D u r a tio n o f e x p e r im e n ts . E x p e r im e n ta l r e s u l t s • 1. 2. D i f f e r e n t i a t i o n o f L in e s . (a ) M o r t a lit y i n g e n e r a t io n " J .M (b ) C om parison o f m o r t a l i t y i n g e n e r a t io n (c ) M o r t a lit y i n g e n e r a t io n "K .” (d ) C om parison o f m o r t a l i t y i n g e n e r a t io n A n a ly s is o f d a t a . (a ) " t” t e s t s . (b ) A n a ly s is o f V a r ia n c e . 3. P a t h o lo g y . 1. B reast r e a c tio n . (a ) B r e a s t r e a c t io n on d i f f e r e n t d a y s . G e n e r a tio n J : 6t h d a y , 1 0 th d a y . G e n e r a tio n K: 6t h d a y , 1 0 th d a y . (b ) B r e a st r e a c t i o n o f s u r v iv o r s from 1 4 th t o 2 8 th day* (c ) B reast r e a c tio n in d if f e r e n t l i n e s . G e n e r a tio n J ; G e n e r a tio n K. 2 . M e t a s t a t ic in v o lv e m e n t o f t h e i n t e r n a l o r g a n s . 3 . M e t a s t a t ic in c id e n c e o f i n t e r n a l o r g a n s . D is c u s s io n Summary R e fe r e n c e s A ck n ow led gm en t PRODUCTION At th e tim e t h e p r e s e n t s tu d y was i n i t i a t e d t h e r e e x i s t e d a t t h e U n ite d S t a t e s R e g io n a l P o u lt r y R e se a rc h L a b o r a to r y a number o f in b r e d l i n e s o f W hite L egh orn c h ic k e n s w ith v a r io u s d e g r e e s o f r e s i s t a n c e t o n a t u r a l l y o c c u r r in g ly m p h om ato sis* L ik e w is e , t h e r e e x i s t e d a tumor m a t e r ia l w h ich was o b ta in e d o r i g i n a l l y from a c h ic k e n i n f e c t e d w it h lym p h om ato sis w h ich was c o n t r a c t e d n a t u r a lly * T h is tum or m a t e r ia l a f t e r numerous and fr e q u e n t p a s s a g e s i n v iv o was term ed " S t r a in RPL 1 6 ” and a t th e tim e o f t h i s s t u d y had b een c a r r ie d a p p r o x im a te ly 61 p a s s a g e s * When tumor m a t e r ia l su c h a s RPL 16 was t r a n s p la n t e d i n t o v e r y young c h ic k s o f c e r t a i n in b r e d l i n e s t h e r e w a s, w i t h in a p e r io d o f a few d a y s , a lym phoid grow th r e s p o n s e a t t h e s i t e o f i n o c u l a t i o n and f r e q u e n t ly m e t a s t a s i s t o o th e r p a r t s o f t h e body* Among t h e m ost s u s c e p t i b l e c h ic k s tumor i n j e c t i o n r e s u l t e d i n d e a th e v e n b e fo r e e x t e n s iv e lym p hoid grow th was e v i d e n t • F u r th e r , i t was o b se r v e d t h a t when tumor m a t e r ia l was t r a n s p la n t e d i n t o youn g c h ic k e n s o f o th e r in b r e d l i n e s t h e r e was l i t t l e or no tumor grow th r e s p o n s e and no m o r t a l i t y , t h u s s u g g e s t in g some d e g r e e o f h o s t r e s i s t a n c e t o a s p e c i f i c tumor t r a n s p l a n t . The o b j e c t o f t h e p r e s e n t s tu d y i s an a tte m p t t o d is c o v e r w h eth er c h ic k e n s w it h in c e r t a i n in b r e d l i n e s t h a t h a v e b e e n s e l e c t e d fo r r e s i s t a n c e or s u s c e p t i b i l i t y t o n a t u r a l l y o c c u r r in g ly m p h o m a to sis w i l l show a s im ila r d e g r e e o f r e s i s t a n c e or s u s c e p t i b i l i t y t o t r a n s p l a n t s o f lym p hoid tum ors* An answ er t o t h i s p rob lem i s o f im p o rta n ce t o t h e s tu d y o f ly m p h o m a to sis b e c a u se under t h e p r e s e n t m ethod o f t e s t i n g c h ic k e n s fo r r e s i s t a n c e or s u s c e p t i b i l i t y t o t h e d i s e a s e a lo n g and c o s t l y h o ld in g p e r io d (6 0 0 d a y s ) - 1- is n ecessary. I f a r a p id t e s t fo r r e s i s t a n c e t o lym p h om atosis c o u ld be d e v e lo p e d t h e b r e e d in g program w ould p r o c ee d more r a p i d l y . DESCRIPTION OF LYMHOMATOSIS L ym p h om atosis, t h e d i s e a s e co n cern ed i n t h i s s t u d y , i s a form o f wThe A v ia n L e u k o s is Complex’* w hich i s a t e n t a t i v e te r m in o lo g y s u g g e s te d b y J u n g h e r r , J o h n so n , and D o y le (1 9 4 1 ) t o c o v e r a group o f d i s e a s e s w hich a re p r im a r ily c h a r a c t e r iz e d by autonom ous p r o l i f e r a t i o n o f t h e e s s e n t i a l b lo o d fo rm in g c e l l s * D e fin itio n : L ym phom atosis may be d e fin e d as a n a t u r a lly o c c u r r in g m a lig n a n t d is e a s e o f c h ic k e n s c h a r a c t e r iz e d by t h e fo r m a tio n o f lym phoid tum ors w hich may be found i n n e a r ly a l l t i s s u e s o f t h e b i r d ’ s b o d y . T here a r e fo u r m ain form s o f ly m p h o m a to sis: n e u r a l, o c u la r , v i s c e r a l , and o s t e o p e t r o t i c , a c c o r d in g t o t h e t i s s u e a f f e c t e d * N e u r a l lym p h om atosis or fo w l p a r a l y s i s a t t a c k s , p r im a r ily , young b ir d s and i s c l i n i c a l l y m a n ife s te d b y asym m etric p a r e s i s o f t h e l e g or w in g . O cu lar ly m p h o m a to sis, commonly c a l l e d g r a y e y e , i s m a n ife s t e d by i r r e g u l a r i t y i n t h e p u p i l , g r a d u a l l o s s o f l i g h t accom m odation, and g r e y c o lo r o f th e i r i s * V i s c e r a l lym p h om atosis a f f e c t s t h e la r g e abdom inal organs such a s t h e l i v e r , s p l e e n , and k id n e y s , bu t any organ o f t h e b od y, in c lu d in g t h e s k i n , may be in v o lv e d a t tim e s * U n le s s t h i s form o f t h e d i s e a s e i s accom panied by s y s te m ic d is tu r b a n c e t h e outward s i g n o f t h e d i s e a s e i s in d e fin ite . O s t e o p e t r o t ic ly m p h o m a to sis, o f t e n c a l l e d m arble bone or t h i c k - l e g d i s e a s e , a f f e c t s t h e lo n g bones o f th e s k e l e t o n and i t c a n be d e t e c t e d by c l i n i c a l i n s p e c t i o n . - 2 - E tio lo g y : C r i t i c a l e x p e r im e n ta l s t u d i e s , s u g g e s t t h a t ly m p h o m a to sis i s c a u sed by one o r more v i r u s or v i r u s - l i k e a g e n ts # T h is a g e n t may be p r e s e n t i n c e r t a i n c h ic k em b ryos, n ew ly h a tc h e d c h i c k s , and i n t h e b lo o d and tum or t i s s u e o f a ff e c te d b ir d s . I t i s n a t u r a l l y t r a n s m it t e d e i t h e r th ro u g h t h e e g g or by c o n ta c t# D o y le (1 9 2 7 ) was t h e f i r s t t o s u g g e s t t h a t t h i s d i s e a s e was t r a n s m it t e d by way o f t h e egg# L a t e r , numerous i n v e s t i g a t o r s , W arrack, and D a lli n g ( 1 9 3 2 ) , B i e l y e t a l . ( 1 9 3 2 ) , S e a g e r ( 1 9 3 3 ) , B lakem ore and G lo v e r ( 1 9 3 5 ) , G ibbs ( 1 9 3 6 ) , Tower ( 1 9 3 7 ) , M cC lary and Upp (1 9 3 9 ) and L ee and W ilck e ( 1 9 4 1 ) , p r e s e n te d t h e e v id e n c e t h a t th e eg g and i n f e c t e d b ir d s a re c a r r i e r s o f lym p h o m a to sis# W aters and P r i c k e t t (1 9 4 4 ) em p h asized t h a t i n f e c t e d b ir d s s e r v e a s a means o f d is e a s e t r a n s m i s s i o n . W aters (1 9 4 5 a ) s t a t e d t h a t lym p h om atosis i s t r a n s m it t e d b o th by way o f t h e eg g and by c o n ta c t. The s u b j e c t o f e g g t r a n s m is s io n o f lym p h om atosis h a s r e c e n t l y b e e n r e v ie w e d by C o le (1 9 4 9 ) and by C o t t r a l (1 9 4 9 b )# C o le 's (1 9 4 9 ) i n t e r p r e t a t i o n o f t h e r e s u l t s o f h i s work was t h a t i f e g g borne t r a n s m is s i o n o f a v ia n l e u k o s i s from i n f e c t e d b r e e d e r s d o es o ccu r i t i s n o t o f g r e a t im p o rta n c e i n d e te r m in in g t h e v i a b i l i t y o f t h e progen y# R e c e n t ly , C o t t r a l , B u rm ester, and W aters (1 9 4 9 a ) h a v e d em o n stra ted t h a t c e r t a i n embryos a r e c a r r i e r s o f t h e a g en t o f lym p h om atosis • T hese e x p e r im e n ts a l s o i n d i c a t e t h a t c e r t a i n n o r m a l-a p p e a r in g h en s tr a n s m it ly m p h o m a to sis t o t h e i r p r o g e n y by way o f t h e egg# I t is e q u a ll y e v id e n t t h a t m o d if ic a ­ t i o n s o f th e en viron m en t (H u tt, e t a l # , 1 9 4 4 ) and g e n e t ic r e s i s t a n c e (H u tt, e t a l# 1 9 4 1 , T a y lo r , e t a l • 1 9 4 3 , and W a te r s, 1 9 4 5 b ) , d e te r m in e t o a l a r g e e x t e n t t h e in c id e n c e o f ly m p h o m a to sis. W aters (1 9 4 7 ) and W aters and B yw aters (1 9 4 9 ) c o n c lu d e d t h a t th e in c id e n c e o f lym p h o m a to sis - 3 - i n e x p e r im e n ta l b ir d s i s d ep en d en t t o some e x t e n t on t h e d e g r e e o f e x p o s u r e , age a t tim e o f e x p o s u r e , and g e n e t ic r e s i s t a n c e . The p r e s e n c e o f t h e a v ia n l e u k o s i s a g e n ts i n t h e b lo o d s u g g e s t s t h a t t h e d i s e a s e may be t r a n s m it t e d by b lo o d -s u c k in g p a r a s i t e s . W h ile m ost o f t h e e x p e r im e n ts w ere n e g a t i v e , J o h n so n (1 9 3 7 ) showed t h a t t h e i n f e c t i o n i s m e c h a n ic a lly t r a n s m it t e d by ccsnmon red m ite and fo w l t i c k . H e a l s o p r e s e n t e d t h e id e a t h a t m inor o p e r a tio n s may h ave t h e same e f f e c t . The d i s e a s e h a s b een r e p o r te d from e v e r y m ajor p o u lt r y c o u n tr y i n th e w o r ld . The l o s s e s from t h i s d is e a s e have b een e s t im a te d t o be $ 6 0 ,0 0 0 ,0 0 0 a n n u a lly i n t h e U n ite d S t a t e s . P o u lt r y R e s . L a b ., E a st L a n s in g , M ich . (U* S . D e p t. A g r i c . , R e g io n a l 1 0 th Annual R e p t . 1 9 4 9 ) At p r e s e n t t h e r e a re no known rem ed ies or m ea su res w h ich w i l l p r e v e n t or cu re ly m p h o m a to sis. T here i s , h o w ev er, enough in f o r m a tio n a v a i l a b l e t o show t h a t in n a t e r e s i s t a n c e or s u s c e p t i b i l i t y t o d i s e a s e e x i s t s i n b r e e d s and v a r i e t i e s o f a n im a ls . A c t u a l l y , i t has b e e n d em o n stra ted t h a t s e l e c t i v e b r e e d in g w i l l produ ce c h ic k e n s h i g h ly r e s i s t a n t t o ly m p h o m a to sis . I n h e r it a n c e i n r e l a t i o n t o d i s e a s e U n t i l q u it e r e c e n t l y t h e v a lu e o f i n h e r it e d q u a l i t i e s a s a f a c t o r i n d i s e a s e phenomena was n o t f u l l y a p p r e c ia t e d . The e v id e n c e a c q u ir e d i n p la n t g e n e t i c s s u g g e s te d t h a t th e i n h e r i t e d q u a l i t i e s o f t h e in d i v id u a l sh o u ld be c o n s id e r e d a s an im p o rta n t h o s t v a r ia b le i n v a r io u s d i s e a s e s and s t r u c t u r a l a n o m a lie s . The y e l l o w mouse l e t h a l (C u e n c t, 1 9 0 8 ) , l e t h a l anem ia o f m ice a s s o c i a t e d w it h w h ite s p o t t i n g ( L i t t l e , 1 9 1 5 ) , a c h o n d r o p la s ia i n c a t t l e (C rew , 1 9 2 3 ) , c r y p to r c h id is m i n g o a ts (Lush e t a l , 1 9 3 0 ) and c r y p to r c h id is m i n sw in e (McPhee and B u c k le y , 1 9 3 4 ) , a re a l l s i g n i f i c a n t exam p les o f - 4- t h e e f f e c t o f in h e r it a n c e o f n o n - i n f e c t i o u s d is e a s e c o n d it io n s i n t h e a n im a l kingdom . R e c e n t ly many w ork ers h a v e b een co n c er n e d w ith s t u d i e s on t h e i n h e r i t a n c e o f s u s c e p t i b i l i t y o r r e s i s t a n c e t o i n f e c t i o u s and p a r a s i t i c d is e a s e s . R o b e r ts and Card ( l 9 ^6 ) and R o b e r ts , S e v e r e n s and Card ( l 9 3 9 a «b*) d e m o n str a te d t h e n a t u r a l r e s i s t a n c e o f fo w l t o S a lm o n e lla p u llo r u m . A ls o , t h e e v id e n c e o f g e n e t i c r e s i s t a n c e o f c h ic k e n s t o fo w l ty p h o id was p r e s e n t e d by Lam bert and Knox (1 9 3 2 )* The r e s i s t a n c e o f fo w l t o t h e round worm A s c a r id ia l i n i a t a h a s b een s t u d ie d by A c k e r t (1 9 5 $ ) and h e r e p o r t e d t h a t c e r t a i n b r e e d s o f c h ic k e n s a r e l e s s r e s i s t a n t t o p a r a s i t i c i n f e c t i o n th a n o t h e r s . I n h e r it a n c e i n r e l a t i o n t o tum ors i n m ic e . I n b r e d l i n e s o f m ic e h a v e b een d e v e lo p e d , w hich show d e f i n i t e t e n d e n c ie s tow ard r e s i s t a n c e o r s u s c e p t i b i l i t y t o sp o n ta n e o u s and t r a n s ­ p la n t a b l e tu m o rs. The e x p e r im e n ta l r e s u l t s by Loeb ( 1 9 0 1 ) , U y z ze r ( 1 9 0 7 ) , H aaland ( 1 9 1 1 ) , C uenot ( I 9 O8 ) , M urray ( 1 9 1 1 ) , S ly e ( I 9 I 5 ) (1 9 3 1 )* Lynch (1 9 2 4 , L i t t l e ( 1916 , 1931* an<* 1 9 4 l ) , B it t n e r (193® , and o t h e r s , i n d i c a t e beyond d ou b t t h a t g e n e t i c s p la y s an im p o r ta n t p a r t i n i n f l u e n c i n g n a t u r a l­ l y o c c u r r in g tum ors i n m ic e . I t i s a ls o c o n c lu d e d from t h e work o f Loeb ( 1 9 0 2 ) , L i t t l e (1 9 1 3 )* L i t t l e and T y zzer ( 1 9 1 6 a . b . ) , L i t t l e and J o h n so n ( 1 9 2 2 ) , Loeb and W right ( 1 9 2 7 ) , B it t n e r ( 1 9 3 9 ) , and o t h e r s t h a t t h e i n c i ­ d en ce and d ev elo p m en t o f t r a n s p la n t a b le tum ors a r e p r im a r ily d e p e n d en t upon t h e g e n e t i c c o n s t i t u t i o n o f t h e m ic e . N a t u r a lly o c c u r r in g lym p h om ato sis P ap p en h eim er, Bunn, and Cone (1 9 2 6 ) s t a t e d t h a t t h e i r o b s e r v a t io n s i n d i c a t e d a s i g n i f i c a n t b reed o r v a r i e t y d i f f e r e n c e i n s u s c e p t i b i l i t y t o ~ 5 - fo w l p a r a l y s i s . D oyle ( 1 9 2 7 ), su g g ested , t h a t fo w l p a r a l y s i s may be t r a n s m i t t e d th ro u g h t h e e g g . Asmundson and B ie ly (1 9 5 ^ ) f u r n i s h e d d a t a i n d i c a t i n g t h a t r e s i s t a n t and s u s c e p t i b l e b i r d s d i f f e r by a s i n g l e d o m in a n t g e n e . A d d itio n a l e v id e n c e i n s u p p o r t o f t h i s t h e o r y was f u r n i s h e d by B ie ly , P a lm e r, L e r n e r , and Asmundson (1 9 5 5 )* ^ s h o u ld be n o t e d , h o w ev er, t h a t i n b o th t h e s e e x p e rim e n ts t h e num ber o f b i r d s in v o lv e d was r a t h e r s m a ll. I t was n o t i c e d by K ennard and C h am b erlin (1954-) t h a t th e f l o c k w ith w hich th e y h a d b ee n w o rk in g f o r s e v e r a l y e a r s was m ore r e s i s t a n t t h a n b i r d s w hich w ere p u rc h a s e d fro m n o n - a f f e c t e d f l o c k s and r e a r e d w ith t h e i r own b i r d s . G ild o w , W illia m s , and Lampman (1 9 5 ^ ) r e p o r t e d t h a t t h e p ro g e n y o f c e r t a i n in d iv id u a .1 h e n s w ere fo u n d t o be m ore r e s i s t a n t th a n t h e p ro g e n y o f o t h e r h e n s . They a l s o found t h a t t h e d i s e a s e d e v e lo p e d m ore e x t e n s i v e l y among t h e p ro g e n y o f a f f e c t e d p a r e n t s to c k t h a n among t h e p ro g e n y o f n o n - a f f e c t e d p a r e n t s . Upp and Tower (1 9 5 ^ ) o b s e rv e d i n h e r e n t d i f f e r e n c e s i n r e s i s t a n c e t o fo w l p a r a l y s i s . W ilc k e, L e e , and M urray (19^&) w ere a b le t o d e v e lo p by s e l e c t i o n two g ro u p s o f b i r d s t h a t p ro v e d t o be d i f f e r e n t w ith r e s p e c t to t h e in c id e n c e o f fo w l p a r a l y s i s i n t h e i r o f f s p r i n g . M arb le (1 9 5 9 ) o b s e rv e d m arked f a m ily d i f f e r e n c e s i n t h e e x t e n t t o w hich r e s i s t a n c e t o fo w l p a r a l y s i s i s d ev elo p e d by s e l e c t i o n m e th o d s. H u t t , C o le and B ru ck n er ( i p 4 l ) came t o t h e c o n c lu s io n a f t e r t h r e e y e a r s o f b r e e d in g to i n c r e a s e v i a b i l i t y . A s i g n i f i c a n t f a c t em erg in g fro m t h e w ork o f H u tt e t a l . (1 9 4 1 , 194-7) i s t h a t by c a r e f u l s e l e c t i o n - 6 - o f b r e e d e r s , b a s e d upon p ro g e n y t e s t , i t was p o s s i b l e t o d e v e lo p l i n e s o f c h ic k e n s t h a t w ere r e s i s t a n t and o t h e r s t h a t w ere s u s c e p t i b l e t o ly m p h o m a to sis . A f t e r e i g h t y e a r s o f p ro g e n y t e s t s e l e c t i o n , T a y lo r e t a l . (1 9 4 5 ) a l s o d e v e lo p e d r e l a t i v e l y r e s i s t a n t and s u s c e p t i b l e l i n e s o f c h ic k e n s w ith r e s p e c t t o ly m p h o m a to sis. S t a r t i n g w ith 9 s t r a i n s o f "White L e g h o rn s , W aters and P r i c k e t t (1 9 4 4 ) d e v e lo p e d i n b r e d l i n e s t h a t e v e n t u a l l y showed s i g n i f i c a n t d i f f e r e n c e s i n t h e i r r e s i s t a n c e t o ly m p h o m a to sis. The d a t a i n t a b l e 1 show t h a t s e l e c ­ t i o n s and i n b r e e d i n g w ere v e r y u s e f u l t o o l s i n s e g r e g a ti n g t h e l i n e s i n t o s u s c e p t i b l e and r e s i s t a n t p o p u l a t i o n s . The cfefca a l s o show t h a t g e n e t ic s e l e c t i o n w ith in b r e e d in g h a s d e c r e a s e d t h e in c id e n c e o f ly m p h o m ato sis i n c e r t a i n l i n e s , w h ile i n o t h e r l i n e s t h e r e h a s b e e n a n i n c r e a s e . W a te rs (1 9 4 5 a ) a c c o u n te d f o r th e p r o g r e s s i v e i n c r e a s e and d e c r e a s e i n t h e in c i d e n c e o f ly m p h o m a to sis a s due to d e f i n i t e s e g r e g a t i o n o f g e n e s f o r r e s i s t a n c e o r f o r s u s c e p t i b i l i t y to ly m p h o m ato sis. E x p e r im e n ta lly t r a n s p l a n t e d ly m phom atosis A lth o u g h a g r e a t d e a l h a s b een r e p o r t e d on t h e t r a n s m i s s i o n o f ly m p h o m a to sis, v e r y l i t t l e w ork h a s b e e n done on t h e r e l a t i o n o f i n h e r i t a n c e t o e x p e r im e n ta lly t r a n s p l a n t e d ly m p h o m a to sis. C ole (1 9 4 1 ) c o n d u c te d e x p e rim e n ts t o d e te r m in e th e e x te n t t o w hich r e s i s t a n c e t o t r a n s m i s s i b l e sarco m a ( J u n g h e r r , 1957) c o u ld be i n c r e a s e d by g e n e t i c s e le c tio n . I n C o le ’ s s tu d y t h e p r i n c i p l e o f p ro g e n y t e s t v^as u s e d and a t o t a l o f 2 676 p e d ig r e e d c h ic k s w ere in o c u la t e d w ith t h e n e o p la s m . The r e s u l t s showed t h a t b re e d in g f o r s u s c e p t i b i l i t y w i t h i n h i s s u s c e p t i b l e l i n e s was n o t s u c c e s s f u l . On t h e o t h e r h a n d , h e su c c e e d e d i n i n c r e a s i n g t h e r e s i s t a n c e o f h i s r e s i s t a n t l i n e s by s e l e c t i o n . - 7 - CP rO ON On KN KN o- OJ CVI KN a O o o p. p. 3 © ON H •H O Bn KN UN cvi vO 3 © e 43 Pi © ON at $ •iH CU| © P ON Pi rH xs © faoco © _rt © I O P. © P-l P •H ON KN ON rH p o I CO a ION ^VJ C "pt nQ KN rH UN co KN r- KN 01 P. o 4 ^ rH © © i-H Pi © O CM © o i—i m pq < Eh o o CVJ U © « © P* © CO $ r'S XJ CO *H *-3 1 0 , 1 1 , and 15 w ere i n o c u l a t e d w ith a c e l l - f r e e f i l t r a t e o f tumor s t r a i n RPL 1 8 . be t h e m ost s u s c e p t i b l e . L in e 15 was fo u n d t o The o th e r l i n e s i n o r d e r o f t h e i r in c r e a s e d s u s c e p t i b i l i t y to t h e i n o c u l a t i o n w ere r a te d a s f o l l o w s : l i n e s 6 , 1 1 , 1 0 , and 7 . F u rth e r work n eed s t o be done b e fo r e t h e s e r e s u l t s can be c o r r e l a t e d to t h e n a t u r a lly o c c u r r in g d i s e a s e . MATERIALS AM) METHODS S o u rc e o f m a t e r ia l f o r in ocu lu m The tumor m a t e r ia l u se d a s an in o cu lu m i n t h i s s tu d y was o b t a in e d from p e c t o r a l m u sc le lym phoid tum ors t h a t were grown on b ir d s o f s u s c e p t i b l e in b r e d l i n e 15 o f t h e U . S . R e g io n a l P o u lt r y R esea rch L a b o r a to r y f l o c k . T h is tumor s t r a i n h a s b een d e s ig n a t e d a s RPL 1 6 . The tum or s t r a i n o r i g i n a t e d n a t u r a l l y from lym phom atous l i v e r o f t h e b ir d E1121A. T h is b ir d was n o t in o c u la t e d and had n ev er b een i n c o n t a c t w ith in o c u la t e d sto c k . The o r i g i n a l l i v e r t i s s u e was in o c u la t e d in t o c h i c k s . When t h e tumor d e v e lo p e d i n t h e b r e a s t m u sc le o f t h e in o c u la t e d c h ic k s , s u i t a b l e d o n o r s w ith la r g e tu m ors w ere s e l e c t e d t o f u r n is h m a t e r ia l to p r o p a g a te t h e tumor i n o t h e r s u s c e p t i b l e l i n e 1 5 c h i c k s . 19^ 5)* (B u rm ester and P r i c k e t t , T hus, t h e tumor was c a r r ie d a p p r o x im a te ly th r o u g h 61 s e r i a l p a s s a g e s . At t h e 6 1 s t p a s s a g e a l a r g e number o f c h ic k e n s w ere in o c u la t e d w it h s t r a i n RPL 1 6 and t h e r e s u l t i n g tum ors w ere h a r v e s t e d when t h e y had reached a s u ita b le s iz e * The tu m ors w ere a s c e p t i c a l l y removed from t h e s e b i r d s , s c i s s o r s m in ce d , and t h e n p o o le d i n a la r g e c o n t a in e r f o r th o r o u g h m ix in g * Sam ples o f t h e m ixed tum or t i s s u e w ere t h e n removed and p la c e d i n s t e r i l e p y r e x g l a s s and t h e t u b e s w ere flam ed and s e a l e d . The t u b e s c o n t a in in g t h e c o l l e c t e d tu m or t i s s u e w ere t h e n fr o z e n s lo w ly by im m ersing t h e t u b e s i n 95 p e r c e n t e t h y l a lc o h o l and t h e te m p e r a tu r e was lo w e r e d a p p r o x im a te ly 1°C p er m in u te by th e a d d it io n o f p i e c e s o f d ry ic e (CO2 ) • A f t e r f r e e z i n g , t h e tu b e s w ere t r a n s f e r r e d and s t o r e d i n a CO2 i c e b ox h a v in g a te m p e r a tu r e ra n g e o f -6 5 t o -7 6 ° C . T h is p ro ced u re p r o v id e d a so u r c e o f in o c u l a t h a t c o u ld be c o n s id e r e d a s n e a r ly u n ifo r m . P r e p a r a tio n o f in o cu lu m The f r o z e n tumor sam p les w ere thaw ed j u s t b e fo r e i n o c u l a t i o n by p la c in g th e s e a le d tu b e s under ru n n in g t a p w a te r . The t u b e s w ere broken and t h e m a t e r ia l was m inced i n a t i s s u e m in cer t o s u b d iv id e th e t i s s u e i n t o a f i n e p a s t e - l i k e m a t e r ia l* The m inced m a t e r ia l was w eig h ed and th e n d i l u t e d w ith 0 .8 5 p e r c e n t NaCL s o l u t i o n i n th e p r o p o r tio n o f 1 : 9 . The b u lk o f t h e r e s u l t i n g c e l l s u s p e n s io n was f i l t e r e d th r o u g h a l a y e r o f s t e r iliz e d ch eese c lo t h . B ir d s o f l i n e 15 w ere u sed fo r s e r i a l p a s s a g e o f t h e tumor s t r a i n b e c a u se t h e y w ere found t o be h i g h l y s u s c e p t i b l e t o b o th n a t u r a l i n f e c t i o n w it h ly m p h o m a to sis and t o t r a n s p la n t e d lym phoid tu m o r s . The tum ors were h a r v e s t e d from a la r g e number o f b i r d s , p o o le d and s to r e d i n t h e manner d e s c r ib e d t o p r o v id e a more u n ifo rm s o u r c e o f in o c u la t h a t c o u ld be u sed - 11 - a t v a r io u s p e r io d s o f tim e d u r in g t h e c o u r s e o f t h e e x p e r im e n ts . A s e p t ic p r e c a u t io n s w ere u s e d to p r e v e n t c o n ta m in a tio n i n o b t a in in g t h e m a t e r ia l and i n p r e p a r in g i t f o r i n o c u l a t i o n . A d m in is tr a tio n o f in ocu lu m The sta n d a r d d o se o f in o cu lu m i n a l l o f t h e e x p e r im e n ts wa 3 0 .1 c c . The r o u te w hich was u se d i n a l l c a s e s was t h e r i g h t p e c t o r a l m u s c le . The i n o c u l a t i o n was made i n t o t h e b r e a s t w ith a 20 gauge n e e d le a tt a c h e d t o a 1 c c . t u b e r c u li n s y r in g e . The n e e d le was i n s e r t e d i n t o t h e p e c t o r a l m u s c le a t t h e a n t e r io r p a r t o f t h e b r e a s t . T h is te c h n iq u e le s s e n e d t h e d an ger o f a c c i d e n t a l i n t r a p e r i t o n e a l i n o c u l a t i o n . R e c ip ie n t s The c h ic k e n s u se d i n t h i s s tu d y were a l l W hite L eghorns and c o n s is t e d o f b o th m a le s and f e m a le s . A lt o g e t h e r , e i g h t d i f f e r e n t in b r e d l i n e s o f c h ic k e n s , r e p r e s e n t in g two c o n s e c u t iv e g e n e r a t io n s , d e s ig n a te d a s nJ" and MKn were in v o l v e d . The h i s t o r y o f t h e e i g h t in b r e d l i n e s , d e s ig n a t e d a s l i n e s 4 , 6 , 7* 9* 10* H * 1 4 , &ud 19* u se d i n t h i s s tu d y i s d e s c r ib e d by W aters (1945* b ) . Of t h e e i g h t l i n e s u s e d , two h a v e b een c l a s s i f i e d a s r e s i s t a n t and s i x a s s u s c e p t i b l e . C h ick s w ere h a tc h e d a p p r o x im a te ly e v e r y two w eeks s t a r t i n g i n S ep tem b er, 1 9 4 8 , and c o n tin u in g u n t i l J a n u a ry , 1949* o f age. A ll c h ic k s were h e ld i n b a t t e r y b r o o d e r s u n t i l t h e y w ere 21 d ays As p r e v i o u s l y d e s c r ib e d a l l c h ic k e n s , a t 21 d a y s o f a g e , r e c e iv e d 0 .1 c c . o f tumor m a t e r ia l i n t h e r i g h t p e c t o r a l m u s c le . o b s e r v e d f o r a 28 day p e r io d a f t e r i n o c u l a t i o n . s u r v iv o r s w ere k i l l e d and exam in ed . A ll c h ic k e n s w ere On t h e 2 8 th day a l l D uring t h e p e r io d th e c h ic k e n s were u n d er o b s e r v a t io n t h e y w ere h e ld i n s p e c i a l h o ld in g b a t t e r i e s and i s o l a t e d from a l l o th e r c h ic k e n s . Dead b ir d s w ere removed s e v e r a l tim e s d a i l y and a l l n e c e s s a r y in f o r m a tio n c o n c e r n in g t h e b ir d was e n te r e d on a ca rd t h a t - 12 - acco m p an ied t h e b i r d t o t h e a u to p s y room . The dead b i r d s w ere r e f r i g e r a t e d u n t i l p o s t m ortem e x a m in a tio n s w ere p e rfo rm e d . T a b le 2 shows t h e t o t a l number o f c h ic k e n s o f b o th s e x e s u s e d i n t h i s e x p e rim e n t a s w e ll a s t h e number o f b i r d s from each l i n e r e p r e s e n t e d . The l i n e s r e p r e s e n t e d i n t h i s e x p e rim e n t w ere d e v e lo p e d i n th e l a b o r a t o r y th r o u g h s e l e c t i v e i n b r e e d in g f o r r e s i s t a n c e o r s u s c e p t i b i l i t y t o n a t u r a l l y o c c u r r i n g ly m p h o m a to sis. (W a te rs , 194-5 a ) TABLE 2 G e n e r a tio n J Fem ale M ale L in e T o ta l M ale G e n e r a tio n K Fem ale T o ta l J and K T o ta l - 61 78 159 159 4 - 6 144 142 286 146 182 52 8 614 7 22 16 58 92 98 190 228 9 25 55 56 76 110 186 242 10 66 64 150 - - - 150 11 61 60 121 27 57 64 I 85 14 55 71 62 60 122 195 63. . . 54 117 296 619 1146 2027 1 5 ........ T o ta l - 56 108 _ 71 459 422 179 . 881 527 O o l l e c t i o n o f d a ta T h ro u g h o u t t h e e n t i r e w ork each i n d i v i d u a l was exam ined f o r c l i n i c a l m a n i f e s t a t i o n o f tu m o rs a t t h e s i t e o f i n o c u l a t i o n on t h e s i x t h , t e n t h , f o u r ­ t e e n t h , s e v e n te e n th , t w e n t y - f i r s t , and tw e n ty - e ig h th day o f s u r v i v a l . A m ethod f o r e v a l u a t i n g t h e gro w th o f t h e tu m o r i n t h e "beast m u sc le was d e v is e d i n w hich s i x c l a s s e s w ere d e s ig n a te d t o c o v e r t h e r e a c t i o n o b ta in e d i n t h e - 15 - b rea st# No v i s i b l e tu m or was s c o r e d i n C la s s 0 , tum ors t h a t w ere n o t more th a n 1 c m ., 2 c m ., 3 c m ., 4 c m ., and more th a n 4 cm. i n d ia m eter w ere grad ed i n C la s s e s 1 , 2 , 3 , 4 , and 5 r e s p e c t i v e l y . The l i v e r was n o t c l a s s i f i e d n u m e r ic a lly , but was c l a s s i f i e d a s 0 , A, or B . No v i s i b l e m e t a s t a s is was i n d ic a t e d by 0 , A I n d ic a t e d a d if f u s e d ty p e o f in v o lv e m e n t w it h no f o c a l tu m o r s, and B i n d ic a t e d a f o c a l ty p e o f in v o lv e m e n t . F req u en t m e t a s t a s i s was o b se rv e d i n e a c h o f t h e f o llo w in g o r g a n s : h e a r t , s p l e e n , k id n e y , p a n c r e a s , p r o v e n t r i c u l u s , g on ad , and p e r ito n e u m , a t t h e tim e o f a u t o p s y . tw o c l a s s e s . i n C la s s 0 . The d a ta c o l l e c t e d was grouped under No v i s i b l e m e t a s t a t ic fo r m a tio n i n ea ch o rg a n was sc o r e d Any m e t a s t a t i c fo r m a tio n fo r e a c h o rg a n was c l a s s i f i e d in C la s s R . The above m ethods o f g r a d in g a re b a sed on p u r e ly a r b it r a r y c o n s id e r a t io n and r e p r e s e n t an e v a lu a t i o n o f t h e i n t e r p l a y b etw een t h e tum or t r a n s p la n t and r e s i s t a n c e o f t h e h o s t . D u r a tio n o f e x p e r im e n ts Each ex p e rim en t l a s t e d 7 w eek s a f t e r t h e h a tc h in g o f t h e c h ic k e n s . The c h ic k e n s t h a t s u r v iv e d w ere k i l l e d on t h e 2 8 th day a f t e r t h e i r i n o c u l a t i o n and t h e y w ere 42 d a y s o f age a t t h e t e r m in a t io n o f t h e e x p e r im e n t. EXPERIMENTAL RESULTS D iffe r e n tia tio n o f lin e s A lt o g e t h e r 2027 p e d ig r e e d W hite L egh orn c h ic k s w ere in o c u la t e d w it h tu m or m a t e r ia l from s t r a i n RPL 1 6 . E ig h t d i f f e r e n t in b r e d l i n e s o f c h ic k e n s r e p r e s e n t in g two c o n s e c u t iv e g e n e r a t io n s d e s ig n a te d a s "Jrt - 14 - and "Kw -were in v o lv e d i n t h e s e s t u d ie s * A l l c h ic k e n s w ere h a tch ed d u r in g t h e l a t t e r p a r t o f 1 S 4 8 . The a v e r a g e p e r c e n ta g e o f m o r t a l i t y f o r e a ch o f t h e in b r e d l i n e s o f c h ic k e n s and fo r e a c h g e n e r a t io n i s shown i n t a b l e 3* "J” g e n e r a t io n c h ic k e n s t h e lo w e s t m o r t a l i t y was in l i n e Among t h e 6 , w h ile t h e h ig h e s t m o r t a lit y was found among t h e c h ic k e n s o f l i n e s 7 and 15* For t h e "K” p o p u la t io n t h e lo w e s t m o r t a l i t y was i n l i n e s 4 and 6 w h ile t h e h ig h e s t m o r t a l i t y was among t h e c h ic k e n s o f l i n e 7 and 1 5 . T here w ere no in b r e d l i n e 4 c h ic k s a v a i l a b l e f o r s tu d y i n t h e MJ W g e n e r a t io n and no in b r e d l i n e 10 c h ic k s a v a i l a b l e i n t h e "K" g e n e r a t io n . A g r a p h ic p r e s e n t a t io n o f th e r e s u l t s shown i n t a b l e 3 i s g iv e n i n f i gure 1 • TABLE 3 P e r c e n ta g e o f M o r t a lit y 28 Days a f t e r I n o c u la t io n among Two D i f f e r e n t G e n e r a tio n s o f In b red C h ick en s I n o c u la t e d w ith Tumor S t r a i n RPL 1 6 . In b red lin e ............. ^ ' T r G e n e r a tio n N o. P ercen t N o. o f b i rds dead dead "v™- G e n e r a tio n N o. N o. o f P ercent dead dead b ir d s .......................... 4 - - - 13 9 32 23 .0 6 286 69 2 4 .1 328 98 2 9 .8 7 38 37 9 7 .4 19 0 189 9 9 .5 9 56 41 7 3 .2 186 128 6 8 .8 10 130 93 7 1 .5 - - 11 121 81 6 6 .9 64 25 3 9 .0 14 71 40 5 6 .3 122 57 4 6 .7 15 179 173 9 6 .6 117 113 9 6 .5 - 15 - - oo E J 90 80 70 60 50 40 30 20 10 O 4 6 7 9 10 LINES Comparison of the total mortality rates for both genera tions in the various fines. 15 - M o r t a lit y i n G e n e r a tio n nJ w The d a ta i n t a b l e 5 show t h a t i n g e n e r a t io n "J 0 t h e l i n e s f o l l o w t h e o r d e r o f 7» 1 5 j 9* 1 0 5 1 1 , 1 4 , ^ r e s p e c t i v e l y , r a n g in g from t h e s u s c e p tib le to th e r e s is t a n t l i n e . As c a n be n o t ic e d from t h e d a ta i n t a b l e 4 , t h e r e i s a v a r i a t i o n b etw een t h e l i n e s f o r t h e d i s t r i b u t i o n o f t h e d e a th r a t e d u r in g t h e p e r io d o f t h e e x p e r im e n t. E x te n d in g o v e r a p e r io d o f 28 d ays t h e c h ic k e n s o f l i n e 6 showed t h e lo w e s t t o t a l m o r ta lity . T hroughout t h e 2 8 -d a y p e r io d a fe w c h ic k e n s i n l i n e 6 d ie d each day w ith t h e h i g h e s t m o r t a l i t y o c c u r r in g on t h e 12 th and 20 t h day fo llo w in g in o c u la t io n . The m o r t a l i t y o f l i n e 7 ( t a b l e 4 ) shows two p ea k s w hich a r e o n t h e 1 2 - 1 4 t h and on t h e 2 0 -2 5 r d d a y s . At t h e f i r s t p e r io d 1 8 . 4 l p e r c e n t o f t h e c h ic k e n s d ie d w h ile i n t h e l a t t e r p e r io d 52*59 p e r c e n t d ie d . O nly one c h ic k e n i n l i n e 7 s u r v iv e d t o th e end o f t h e e x p e r im e n t. The maximum p e r c e n t a g e o f m o r t a l i t y rea c h e d i n l i n e 9 was 1 0 .7 0 p e r c e n t on t h e l^ t h day a f t e r i n o c u l a t i o n . The h i g h e s t p e r c e n ta g e o f d e a th s (4^*52 p e r c e n t ) o c c u r r e d b etw een t h e 12 t h and 17 th day and 17*84 p e r c e n t o f t h e c h ic k e n s d ie d b etw een th e l p t h and 22 nd d a y s . Of t h e 7 1 * 5 p e r c e n t d e a th s i n l i n e 1 0 , 2 ^ .0 6 p e r c e n t and 29*21 p e r ­ c e n t o f t h e d e a th s o c c u r r e d o n t h e l l t h - 1 4 t h and t h e I 6 t h - 20 t h d a y s , r e s p e c tiv e ly . The a v e r a g e m o r t a li t y w it h in t h e s e two p e r io d s was app rox­ im a t e ly t h e same f o r each d a y . I n t h e f i r s t p e r io d t h e a v e r a g e m o r t a l i t y was 5*79 p e r c e n t p e r day and i n t h e seco n d i t was 5*99 p e r c e n t . The m o r t a l i t y i n l i n e 11 i s c h a r a c t e r iz e d by a s e r i e s o f s m a ll p e a k s , none o f w hich a r e s i g n i f i c a n t l y h ig h when compared t o th e a v e r a g e m o r t a l i t y o f 5*95 p e r c e n t . - 17- Kr « UN OJ KN CVJ CVJ cvj ON sO • o $• o 3• fH $• o &• o -d " t—i CVI * OJ s• cvj o LTV • ©• P Percentage rH * CVI iH K\ fH rH iH O rH KN UN • rH UN NO • rH 1 1 C— O • KN UN ON • -= t 1 1 O KN • CVI CO C"• UN 3• UN UN 9 O o KN 3« rH KN CVJ • CVJ 3• R• • C*- 1 CVJ CO 9 C"- o rH CO • CVJ t• rH r KN 1 K• KN • CVJ o rH • rH &• LTV iH KN KN KN NO • CVJ LTN KN • UN O O • O rH KN NO CO C"- UN tH • OJ OJ rH nO -=»t -3 - • ©• nO c~- CO KN • UN O KN • rH CVJ ON • CO €• KN NO • CVJ 3• fH nO • -= t KN sO o o-• o 3• * rH V0 • o 3* fH ♦ CVJ rH 3 OJ rH KN KN • KN UN On 81* 1 UN UN • O O KN NO CVJ rH CO rH «H OJ CO OJ iH KN iH * rH CO • CVI CVI o • • L f\ KN • UN NO • -= t UN ON • -d - UN i—I • NO UN NO • iH 3• 00 CVI ON CVJ GO • 39 kn ©• o KN NO • CVJ c— rH -3* o c*— • OJ pH o • UN NO CVJ • c- o UN • KN CVI O rH UN CVJ c vj CVJ CVJ NO 1 i KN UN • tH 1 9 -d - o 1 CO C"• fH 1 1 1 NO f- O i—1 rH tH -d o • o- 1 • rH KN tH • -= f KN t3* 'S'• 1 O rH 1 3• o rH • UN KN CO KN • UN • UN KN &N ON CO • tN- 3* r- cvj lO * S'• CVJ • CM • CTN CD £f•N ON LTN 1 cvj O CO O a vO fH 1 KN GO tH C-• •H m o 5• rH KN • kn c— pH NO c• t• o rH u 3 % 1 UN KN • LTV CVJ CO fH KN vO 3• CVJ 00 1 O KN • CVJ O -3* • rH f• rH 3• • 3• rH 00 1 nn o M ortality for the Various Days after Inoculation Lines of Generation "J" in the D if fe r e n t ■3 KN • O o• UN 1 03 1 t-1 O n rH UN rH On t h e 1 0 th d ay 4*22 p e r c e n t d ie d i n l i n e 14 and t h e d e a th r a t e i n c r e a s e d u n t i l i t re a c h e d 12*70 p e r c e n t on t h e 15 t h d a y . I t d e crease d o n t h e 1 4 th and 15 t h and 1 6 th d ay s and t h e n in c r e a s e d a l i t t l e o n t h e 17 t h and 1 8 th and c o n tin u e d d e c r e a s in g on th e 2 1 s t , 22 n d , and 2 ^ rd days* L in e 15 w hich i s h i g h l y s u s c e p t i b l e a ls o shows two p e r io d s i n w hich t h e m o r t a l i t y r a t e was h ig h * On t h e 10 t h day 5*02 p e r c e n t d ie d , w hich i s t h e h i g h e s t d e a th r a t e a t t a i n e d i n an y l i n e on t h a t day* On t h e 1 1 th and 1 2 th d ay s 25*10 p e r c e n t and 25*50 p e r c e n t d i e d . t h e 15 t h d ay a t o t a l o f 67 p e r c e n t h ad d ie d . By M o r t a l i t y d ropped s h a r p ly t o 0 * 5 5 p e r c e n t o n t h e 1 6 th day t h e n i n c r e a s e d t o 1 0 .6 0 p e r c e n t on t h e l p t h d ay and t h e n d e c r e a s e d t o 7*82, 5 *5 8 , 2 . 2 5 * an(* 0 - 5 5 p e r c e n t on t h e 2 0 th , 2 1 s t , 2 2 n d , and 2 5 rd d ays r e s p e c t i v e l y . The t o t a l m o r t a l i t y i n l i n e 15 was 9 6 . 01 p e r c e n t . C o m p ariso ns o f M o r t a l i t y i n G e n e r a tio n The f i r s t d e a th s i n t h e " J ” g e n e r a t i o n o c c u r re d on t h e 9*th d ay . L in e 6 and l i n e 9 each showed more m o r t a l i t y . On t h e 10 t h day d e a th s o c c u r r e d among t h e c h ic k e n s o f l i n e 6 , 1 0 , 1 4 , and 15* On t h e 1 1 t h day c o n s id e r a b le v a r i a t i o n o c c u r re d i n t h e p e r c e n ta g e o f d e a th s o f t h e c h ic k e n s i n t h e v a r io u s l i n e s . m o r t a l i t y , show ing 25*1 p e r c e n t . L in e 15 had t h e h i g h e s t An i n c r e a s i n g p e r c e n ta g e o f m o r t a l i t y o c c u r r e d i n t h e o t h e r l i n e s e x c e p t f o r l i n e 9 w hich showed no d e a th s . S t a r t i n g w ith t h e 1 2 th day a l l l i n e s showed some m o r t a l i t y w ith t h e h i g h e s t p e r c e n ta g e a g a in p r e s e n t i n l i n e 1 5 * On t h e 1 4 th day a l l l i n e s showed a m o r t a l i t y o f l e s s th a n 5 p e r c e n t . D u rin g t h e f i r s t 14 d ay s l i n e 15 showed a m o r t a l i t y o f a p p r o x im a te ly 6 6 p e r c e n t w h ile l i n e 6 h ad l e s s th a n 9 p e r c e n t m o r t a l i t y . - 19 - A co m p a riso n of' t h e d u r in g t h e 28 - d a y p e r i o d m o r t a lit y f o r a l l l i n e s i n t h e "J" in d ic a te s th a t lin e s 7 g e n e r a t io n 15 w ere t h e m o st s u s c e p t i b l e w h ile l i n e 6 was t h e m o st r e s i s t a n t . I n l i n e 15 m o st o f t h e c h ic k e n s d ie d p r i o r t o 15 days o f ag e w hereas i n l i n e 7 m o st o f t h e d e a th s o c c u r r e d a f t e r t h e 15 - d a y p e r i o d . M o r t a l i t y i n G -en e ra tio n MKn As i s re s is ta n t. shown i n t a b l e 5* l i n e s 4 and 6 p ro v e d t o be t h e m ost D eath from t h e tu m o rs w ere 99*5 p e r c e n t and 9 6 .6 p e r c e n t i n l i n e s 7 end 15 r e s p e c t i v e l y . S t a r t i n g from t h e m o st s u s c e p t i b l e t o th e m o st r e s i s t a n t , t h e l i n e s c a n be a r ra n g e d a s 7 * 1 5 * 9 * 1 4 , 1 1 , 6 , 4 . L in e 4 showed 5*59 p e r c e n t m o r t a l i t y on t h e 12 t h d ay , th e n i t dropped and a g a in i n c r e a s e d on t h e 1 6 th d a y . B etw een t h e 1 6 th and 2 1 s t d ay , 15*91 p e r c e n t o f t h e b i r d s d ie d . I n l i n e 6 th e h i g h e s t m o r t a l i t y (5*79 p e r c e n t ) was re a c h e d on t h e 1 2 th d a y , and 19*49 p e r c e n t d ie d b etw een t h e 1 1 th and 1 4 th d a y s . M o rta lity was l e s s t h a n 1 p e r c e n t d u rin g th e re m a in in g d ays o f t h e e x p e rim e n t e x c e p t on t h e 1 5 t h and 20 t h when i t was I .5 2 p e r c e n t . Among t h e c h ic k e n s o f l i n e 7 w hich w ere h i g h l y s u s c e p t i b l e to tum or i n o c u l a t i o n , 9 9 - 5 p e r c e n t d ie d p r i o r t o t h e 2 5 t h day a f t e r i n o c u l a t i o n . The g r e a t e s t l o s s e s o c c u r r e d b etw een t h e 1 1 th and 1 7 th day and a c c o u n te d f o r 9 4 p e r c e n t o f t h e t o t a l d e a th s . L in e 9 showed a 55*84 p e r c e n t m o r t a l i t y b etw een th e 1 1 th and 1 7 th d a y s . A f te r t h e 1 7 t h day t h e p e r c e n ta g e o f m o r t a l i t y ra n g e d b etw een 5*2 p e r c e n t and O.5 p e r c e n t . L in e 11 o f t h e nKu g e n e r a tio n p erfo rm e d s i m i l a r l y to w hat i t d id i n t h e “J 0 g e n e r a t i o n . The m o r t a l i t y a g a in was c h a r a c t e r i s e d by a s e r i e s - 20 - of* sm a ll p e a k s none o f w h ich w ere s i g n i f i c a n t l y h ig h when com pared to t h e o v e r a l l a v e ra g e d e a th r a t e o f 5*2 p e r c e n t p e r day f o r t h i s l i n e . L in e 14 r e a c h e d i t s h i g h e s t m o r t a l i t y on t h e 1 1 th , 1 2 th , and 15 t h d a y s and a c c o u n te d f o r 2 8 .1 9 p e r c e n t . M ost o f t h e m o r t a l i t y i n l i n e 15 o c c u r re d b e f o r e t h e 1 5 th day a f t e r i n o c u l a t i o n and d u r in g t h i s p e r io d 95*1 p e r c e n t o f t h e b i r d s d i e d . L in e 15 i n t h e nKn g e n e r a t io n d i f f e r s from l i n e 15 i n t h e "J" g e n e r a t i o n i n t h a t t h e m o r t a l i t y was h i g h e r (55*3 p e r c e n t ) i n th e fo rm e r. T h is p e a k m o r t a l i t y o c c u r r e d on th e l 4 t h d ay , w h ile i n th e “J 11 g e n e r a t i o n t h e m o r t a l i t y was h i g h e r o n t h e 1 1 th day (2 5 p e r c e n t ) . F u r th e r m o r e , no s e c o n d a ry r i s e i n m o r t a l i t y o c c u r re d d u rin g th e l a t t e r p e r i o d i n t h e ,lKH g e n e r a t i o n . C om parisons o f M o r t a l i t y L in e s i n G e n e r a tio n BKW F o llo w in g i n o c u l a t i o n t h e f i r s t d e a th s i n t h e "K" g e n e r a ti o n o c c u r re d o n t h e 1 0 th d a y . The h i g h e s t p e r c e n ta g e o f d e a th s o c c u rre d among t h e b i r d s o f l i n e 15 w h ile l i n e s 11 and 14 show no d e a t h s . (T a b le 5 ) On t h e 1 2 th day a f t e r i n o c u l a t i o n t h e m o r t a l i t y p i c t u r e changes ra p id ly . I n l i n e 1 5 , 25*9 p e r c e n t o f th e b i r d s w ere dead and a l l o t h e r l i n e s e x c e p t l i n e 11 showed i n c r e a s e s i n m o r t a l i t y . On t h e 1 5 t h day l i n e s 7» 14, and 15 a l l showed a h ig h m o r t a l i t y w h ile l i n e s 4 , 6, an d 9 showed a d e c r e a s e i n m o r t a l i t y o v e r th e p r e v io u s day. No d e a th s h a d a s y e t o c c u r re d i n l i n e 1 1 . S t a r t i n g w ith t h e 15 t h day n e a r l y a l l l i n e s showed p e r c e n ta g e o f m o r t a l i t y o v e r t h e h ig h o f p r e v io u s p e r i o d s . a d ec re a se in A f te r t h e 1 5 t h day none o f t h e l i n e s ex ceed ed a m o r t a l i t y o f 8 p e r c e n t and d u r in g t h i s l a t t e r p e r io d l i n e 11 u s u a l l y h ad t h e h i g h e s t p e r c e n ta g e o f m o r t a l i t y . - 21 - Cumulative Percentage Mortality for Succeeding Days after Inoculation ( Generation J ) 100 90 80 70 60 o 50 5? 40 30 20 IO II 12 13 14 15 16 17 18 19 20 21 22 23 24 Days after inoculation Cumulative Percentage Mortality for Succeeding Days after Inoculation (Generation K ) 100 90 ax - 80 70 60 50 40 30 20 IO Days after inoculation - 22 - 25 26 27 rH oCVJ • O VO CVJ I tn CVJ 1 Ov • O 1 vO • 1 1 1 o kn • o l$N • 1 1 1 1 i I 1 3 • -J- 1 1 1 1 i—1 CO • rH o 1 3 KN NO • in CO • rH OJ 1 1 rH • kn rH 1 1 cvj in • in • o i & • o in o € o in th e o KN CVJ CVJ CVJ rH b V3 1 CVJ rH • f- © in iH vD <—I • CVJ LT\ • iH KN iH CVJ rH iH iH rH rH ■& • in iH CVJ in • o rH 'O • rH in OJ • NO O • in o 1—I ON • o o SO • o• rH rH vO # rH rH kn rH KN kn NO f t• o rH • CVJ in • o r— rH • 1 i «H o .s • c— rH kn • rH • ON CVJ in rH NO rH o o rH -d - vO C— ON • 1 kn CO • -d - • CVJ o ON • 1 C" • rH UN CO • O o KN • kn rH KN KN o • O CO iH KN kn KN • rH C'kn in kn o 1 3• o • nO i rH rH C-* KN NO • rH 00 • O O CVJ rH i f t• 1 ON C-• in 1 0J in • c^ OJ in • e- On in • m i o kn in NO • CVJ rH 5vT » KN rH 00 • C"- CVJ in • 1—1 &• -3- o &• o ITS rH 00 c • in • NO i3• • * -C f O VO • rH 3* kn O & m rH NO in • rH c*• O o rH CVJ CVJ • rcN nO • CVJ CVJ in • in • rH CVJ in • o ON UN • fH rH nO OJ o KN o* o o KN irv «H M ortality Percentage for the Various Days after Inoculation D ifferent Lines of Generation MKtt o kn rH 10» 11» &2id 14 assum ed a n i n t e r m e d i a t e p o s i t i o n i n r e g a r d t o s u s c e p t i b i l i t y and l i n e s t h e m o st r e s i s t a n t . 6 and 4 w ere fo u n d t o be I n t h e " J ” p o p u la t io n th e c u rv e s i n a l l t h e l i n e s a r e m ore g r a d u a l , b u t i n t h e g e n e r a t io n th e c u rv e s o f th e s u s c e p t i b l e l i n e s r i s e s h a rp ly . I n g e n e r a l , a lth o u g h a co m p ariso n o f th e d a i l y m o r t a l i t y p e r c e n ta g e s f o r t h e v a r i o u s l i n e s o f t h e ttKn and " J " g e n e r a tio n s showed c o n s id e r a b le v a r i a t i o n , t h e c u m u la tiv e m o r t a l i t y p e r c e n ta g e a t t h e end o f t h e e x p e rim e n t was re m a rk a b ly s i m i l a r . A n a ly s is o f D ata The d a t a p r e s e n te d i n t a b l e s 4 and 5 f i g u r e s 1, 2, and 5 show t h a t t h e r e w ere s i g n i f i c a n t d i f f e r e n c e s betw een t h e l i n e s w i t h i n ea c h g e n e r a t i o n i n r e g a r d to p e r c e n ta g e o f m o r t a l i t y ( o r p e r c e n ta g e o f s u r v iv a l) . I n o r d e r t o d e te r m in e w h e th e r o r n o t d i f f e r e n c e s b etw een g e n e r a t i o n s , b e tw ee n s e x e s , and b etw een f a m i l i e s w ith i n l i n e s in f lu e n c e d t h e r e s u l t s , t h e f o llo w in g t e s t s w ere m ade, u s in g t h e p e r c e n ta g e o f s u r v i v o r s : A lit n t e s t was made u s in g l i n e s t h a t had f a m i l i e s common to b o th g e n e ra tio n s . As i s shown i n t a b l e 6 t h e r e was no s i g n i f i c a n t d i f f e r e n c e b etw ee n t h e g e n e r a t i o n s , b etw e en t h e two s e x e s , b etw een fe m a le s i n th e two g e n e r a t i o n s , and b etw een m a le s i n t h e two g e n e r a t i o n s . As a r e s u l t o f n o t f i n d i n g d i f f e r e n c e s b etw een t h e g e n e r a t io n s o r sex es i n s im ila r lin e s (6 , J9 9* 1^> - 24 - 1 5 ) s e x e s and g e n e r a tio n s TABLE 6 ^ t ” T e s t fo r S u r v iv a l P e r c e n ta g e s L in e S ex 6 9 14 15 M ales "J" 86*8 1 8 .2 45 .5 3 .1 M ales "K" 8 6 .4 1 5 .8 6 0 .0 3 .8 n t *i 0 .0 5 9 0 .1 6 5 0 .6 6 5 0 .1 4 6 F em ales MJ t* 8 5 .3 3 3 .3 5 4 .5 0 F em ales "K*1 8 5 .4 2 0 .0 5 0 .0 0 ”t ” 0 .1 3 2 0 .8 5 2 0 .2 0 6 0 M ales "J" and "K" 8 6 .4 1 6 .6 5 2 .5 3 .4 F em ales ttJ*1 and "K11 8 4 .6 2 2 .8 5 2 .5 0 8 5 .2 nt « 0 .2 0 4 - 25 - CO M ales and F em ales ”Ktt 0 5 0 .0 1 8 .1 55 .0 2 .0 0 .5 8 7 0 .3 3 • 8 6 .1 CD M ales and F em ales ”J M 0 CO 0 .6 9 6 * 0 .4 0 0 CO nt n 0 .0 2 w i t h i n f a m i l i e s i n e a c h l i n e w ere com bined and a c h i- s q u a r e t e s t was c o n d u c te d t o s e e i f t h e r e w ere s i g n i f i c a n t d i f f e r e n c e s i n m o r t a l i t y b e tw e e n f a m i l i e s i n t h e same l i n e . D a ta fro m l i n e The m ethod u s e d i s shown i n t a b l e 7* p a r e p r e s e n t e d a s a n exam ple. L in e s t h a t showed s i g n i f i c a n t d i f f e r e n c e s b e tw e e n f a m i l i e s w ere l i n e s 6 , 9 , 1 1 , 1 4 , and 1 5 * F ig u r e s 1 t o 5 g iv e t h e d a t a on t h e d e a th r a t e . From t h e d a ta a s s e m b le d , t h e a v e r a g e s u r v i v a l i n d ay s f o r ea c h l i n e was com puted, u s i n g o n ly t h e b i r d s t h a t d ie d w i t h i n th e e x p e r im e n ta l p e r i o d . B oth t h e " t " t e s t and t h e a n a l y s i s o f v a r ia n c e w ere em ployed t o t e s t t h e d i f f e r e n c e s . 11t w T e s ts T a b le 8 g iv e s t h e r e s u l t s o f a p r im a ry ttt ” t e s t and t h e f o llo w in g o b s e r v a t i o n s w ere m ade: 1. T h e re was a s i g n i f i c a n t d i f f e r e n c e a t t h e 1 p e r c e n t l e v e l b e tw e e n t h e m a le s o f g e n e r a t i o n r,J n and m a le s o f g e n e r a t i o n “K” . 2. th e a t th e 5 p e rc e n t le v e l b etw een fe m a le s o f t h e two g e n e r a t i o n s . 5* nJ n and 4. th e T h e re was a s i g n i f i c a n t d i f f e r e n c e T h e re was no s i g n i f i c a n t d i f f e r e n c e b etw een s e x e s f o r g e n e r a t io n s nKtt co m b in ed . T h e re was a s i g n i f i c a n t d i f f e r e n c e two g e n e r a t i o n s g ro u p in g t h e a t th e 1 p e rc e n t le v e l b etw een s e x e s i n each g e n e r a tio n t o g e t h e r . The same p r o c e d u re was fo llo w e d to f in d o u t i f t h e r e was a d i f f e r e n c e b e tw e e n t h e s e x e s w i t h i n e a c h g e n e r a t i o n . S exes w ith i n g e n e r a t i o n “J 11 d id n o t show an y s i g n i f i c a n t d i f f e r e n c e w h ile t h e r e was a s i g n i f i c a n t d i f f e r e n c e a t t h e 5 p e r c e n t l e v e l b etw een m ales and fe m a le s i n g e n e r a ti o n ttK*. Thus t h e s e x d i f f e r e n c e s n o te d betw een t h e two p o p u la tio n s may be a t t r i b u t e d t o t h e i n f l u e n c e o f g e n e r a t i o n o r t h e i n t e r a c t i o n betw een s e x e s and g e n e r a t i o n s . TABLE 7 X2 T e s t o f F a m ilie s o f L in e 9 G e n e r a tio n s J and K N o. i n F a m ily F a m ily 404 405 8 1 1 7 19 18 17 10 8 12 6 9 12 7 15 7 7 15 25 6 8 ho6 407 409 4 ll 412 415 4 l6 418 419 420 422 425 424 425 427 428 450 431 T o ta l _ P » 68 225 No. L iv in g X 1 10 2 7 5 1 5 2 6 1 2 2 5 5 2 5 5 6 2 4 0 .1 2 5 0 0 .9 0 9 0 0 .2 8 5 6 0 .5 6 7 9 0 .1 6 6 6 0 .0 5 8 8 0 .5 0 0 0 0 .2 5 0 0 0 .5 0 0 0 0*1666 0 .2 2 2 2 0 .1 6 6 6 0 .7 1 4 3 0 .2 0 0 0 0 .2 8 5 6 0 .4 2 8 4 0 .2 3 0 7 0 .2 4 0 0 0 .5 5 5 5 0 .3 0 0 0 = 0.^022 D eg re e s o f fre e d o m - 19 _ = 1 - 0 .5 0 2 2 = 0 .6 9 7 6 q X2 - 4PX - ?£X = P r o d u c ts PX 0 .1 2 5 0 9 .O 9 O9 0 .5 7 1 2 2 .5 7 5 5 0 .5 0 0 0 0 .0 5 8 8 0 .9 0 0 0 0 .5 0 0 0 3 .0 0 0 0 0 .1 6 6 6 0 .4 4 4 4 0 .5 5 5 5 5*5715 0 .6 0 0 0 0 .5 7 1 2 I .2 8 5 6 0 .6 9 2 3 1 .4 4 0 0 0 .6 6 6 6 2 .0 0 0 0 2 9 .2 9 2 7 68 225 P ro b a b ility o f S u r v iv a l P 4 1 .4 5 * * p ? ** S i g n i f i c a n t a t 1 p e r c e n t l e v e l . - 27 - TABLE 8 111° T e s t o n Common F a m ilie s i n “j " and "K11 G e n e r a tio n s U sin g t h e A verage S u r v iv a l Day M ale s G e n e r a tio n " J" : ** M ales G e n e r a tio n nKM : F e m a le s G e n e r a tio n l,J n : F em a le s G e n e r a tio n t = 2 .2 5 t « 0 .6 3 2 t - 5 .5 4 : M a le s G e n e r a tio n MJ U and uKn : F em ales G e n e r a tio n ”J ” and nK,t: G e n e r a tio n 11J 11: G e n e r a tio n llKn : ** S i g n i f i c a n t a t 1 p e r c e n t l e v e l * S ig n ific a n t a t 5 p e rc e n t le v e l - 28 - A n a ly s is o f V a ria n c e The m ethod o f ^ e x p e c te d s u b c la s s n u m b ers” (S n e d e c o r and Cox, 1955) was fo u n d to be m o st a p p r o p r i a t e f o r t h i s a n a l y s i s u n d e r t h e c ir c u m s ta n c e s o f t h e s e e x p e r im e n ts . The d a t a v a r i e d b e c a u s e t h e r e w ere d i s p r o p o r t i o n a t e num bers o f b i r d s i n t h e v a r i o u s f a m i l i e s and a n u n e q u a l se x d i s t r i b u t i o n w i t h i n t h e f a m i l i e s w hich may be a t t r i b u t e d to t h e a c c id e n ts o f s a m p lin g . I t was assum ed t h a t t h e p o p u la tio n from w hich t h i s sam ple was draw n r e a l l y h ad p r o p o r t i o n a t e s u b c la s s num bers. T a b le s 9 a n d 10 show t h e p ro c e d u re o f a n a l y s i s and th e r e s u l t s . I n G e n e r a tio n " J 11 t h e r e was a h i g h l y s i g n i f i c a n t d i f f e r e n c e b etw een s e x e s an d a l s o a l i n e d i f f e r e n c e . I n g e n e r a t io n “K” t h e r e was a h i g h l y s i g n i f i c a n t d i f f e r e n c e b etw een t h e l i n e s . The a n a l y s i s d id n o t show any d i f f e r e n c e b etw een s e x e s i n t h i s g e n e r a ti o n . PATHOLOGY B reast re a c tio n O ut o f a t o t a l o f 2027 b i r d s w hich w ere i n o c u la te d *ri.th t h e lym phoid tum or o n ly t h r e e b i r d s f a i l e d to d ev e lo p breadi tu m o rs . Two o f th e s e b i r d s w ere from r e s i s t a n t l i n e 6 and one was fro m l i n e 11 o f g e n e r a t i o n *\J,f. A f te r t h e g ro w th o f t h e im p la n t had become e s t a b l i s h e d i n some b i r d s t h e tu m o r r e g r e s s e d , i n o t h e r s i t c o n tin u e d to grow, and s t i l l i n o t h e r s i t c o n tin u e d to grow and m e ta s ta s iz e d to t h e v i s c e r a l o r g a n s . The s i z e o f t h e tum or d i f f e r e d and a p p a r e n tl y was r e l a t e d to th e le n g t h o f th e s u rv iv a l p e rio d . I n a l l o f t h e s u s c e p t i b l e b i r d s o f t h e v a r io u s l i n e s th e tum or d e v e lo p e d i n t h e bressfc and m e t a s t a s i s o c c u r re d t o t h e l i v e r , s p le e n , and k i d n e y s . H ow ever, many o f t h e b i r d s , p a r t i c u l a r l y o f l i n e s 7 and 1 5 , - 29 - TABLE 9 A n a ly s is o f V a ria n c e G e n e r a tio n " J" 1. P r im a r y a n a l y s i s o f v a r ia n c e o f m eans o f s u r v i v a l days ( a c t u a l ) S o u rc e o f V a ria tio n D eg rees o f Freedom T o ta l B et\*een s u b c la s s e s (sex lin e ) W ith in s u b c la s s e s 2. Sum o f S q u a re s 332 1 0 , 9 5 2 .8 6 13 6 7 1 .8 6 519 1 0 ,2 8 0 .6 0 Mean S q u are 1 9 .8 0 A n a ly s is o f v a r i a n c e o f m eans o f s u r v i v a l days (m ethod o f e x p e c te d s u b c la s s num ber) S o u rc e o f V a ria tio n Sum o f S q u a re s D eg rees o f Freedom Mean S quare 13 2 ,0 7 1 .0 6 B etw een s e x m eans 1 2 3 2 .1 2 2 5 2 . 12 ** B etw een m eans o f l i n e s 6 1 ,7 5 5 -2 7 289.21** In te ra c tio n 6 1 0 3 .6 7 1 7 .2 3 519 1 0 ,2 8 0 .6 0 1 9 .8 0 B etw een s u b c la s s e s W ith in s u b c la s s e s ** S i g n i f i c a n t a t 1 p e r c e n t l e v e l . - 30 - TABLE 10 A n a ly s is o f V a ria n c e G e n e r a tio n nK" 1. P rim a ry a n a l y s i s o f v a r i a n c e o f means o f s u r v i v a l days ( a c t u a l ) S o u rc e o f V a ria tio n T o ta l B etw een s u b c la s s e s (se x lin e ) W ith in s u b c la s s e s 2. D eg rees o f Freedom Sum o f S q u ares 637 6 , 1 1 1 .5 2 15 1 , 1 9 1 .8 8 624 5 ,9 2 0 .6 4 A n a ly s is o f v a r ia n c e o f m eans o f s u r v i v a l d ay s (m ethod e x p e c te d s u b c la s s num ber) S o u rc e o f V a ria tio n D eg rees o f Feeedom Sum o f S q u ares B etw een s u b c la s s e s 13 1 ,5 8 9 -7 5 B etw een s e x m eans 1 1 .8 8 Betvreen m eans o f l i n e s 6 1 , 2 5 9 .5 4 I n te ra c tio n 6 1 4 8 .5 5 624 5 ,9 2 0 .6 4 W ith in s u b c la s s e s ** S i g n i f i c a n t a t 1 p e r c e n t l e v e l . * S i g n i f i c a n t a t 5 p e r c e n t le v e l# - 31 - Mean S q u are 9 .4 8 of Mean S q u are 1 .8 8 206.59** 24.7 2 * 9 -4 8 d ie d i n a s h o r t tim e so t h a t t h e tum ors d id n o t a t t a i n a g r e a t s i z e * T h u s, t h e s i z e o f th e tum or d i f f e r e d and a p p a r e n tly was r e la t e d t o t h e l e n g t h o f th e s u r v iv a l p e r io d • T h is i s a l s o t r u e fo r t h e b ir d s t h a t l i v e d lo n g e r from t h e s u s c e p t i b l e l i n e s 7 and 15 w h ich u s u a l l y d e v e lo p e d l a r g e r tum or m a sses i n th e b r e a s t and v i s c e r a l o rgan s* E x a m in a tio n o f t h e b ir d s b eg a n on t h e 6 th day a f t e r in o c u l a t i o n b e c a u s e p a s t e x p e r ie n c e had shown t h a t no a p p r e c ia b le b r e a s t tumor would d e v e lo p p r io r t o t h a t t i m e . The d a ta on t h e d e g r e e o f in v o lv em en t o f th e b r e a s t w ere u sed t o com pare t h e v a r io u s l i n e s on th e 6 th and 1 0 th d ays a f t e r i n o c u l a t i o n ( s e e t a b l e s 11 and 1 2 )* The p e r c e n ta g e o f b ir d s t h a t had b r e a s t r e a c t i o n s on t h e s e d ays o f t h o s e t h a t s u r v iv e d ( t a b l e s 15 and 1 6 ) t o th e end o f t h e ex p e rim en t was compared t o th e p e r c e n ta g e o f th e n o n - s u r v iv a ls ( t a b l e s 13 and 1 4 ) t h a t showed a b r e a s t r e a c t io n on t h e s e same d a y s . TABLE 11 The P e r c e n ta g e o f C h ick en s t h a t Showed V i s i b l e B rea st Tumors i n t h e V ariou s In b red L in e s o f G e n e r a tio n on t h e 6 th and 1 0 th Day a f t e r I n o c u l a t i o n . P e r c e n t Showing Tumors on 6 th Day L in e T o t a l No* o f C h ick en s 6 286 2 9 .9 8 9 .8 7 38 7 .9 8 6 .9 9 56 3 5 .8 9 2 .9 10 130 3 3 .9 9 5 .4 11 121 1 9 .1 7 8 .6 14 71 3 6 .7 93 .0 15 179 19*6 9 1 .1 - 32 - P e r c e n t Showing Tumors on 1 0 th Day B r e a s t r e a c t i o n on d i f f e r e n t days . G e n e r a tio n 6 t h Day A co m p a riso n o f t h e l i n e s ( t a b le 1 1 ) showed t h a t t h e r e was no g r e a t d i f f e r e n c e e i t h e r i n t h e p e r c e n ta g e o f b ir d s show ing a b r e a s t tumor or i n t h e d e g r e e o f in v o lv e m e n t i n in d i v id u a l b i r d s . In a l l th e l i n e s , b ir d s t h a t had b r e a s t r e a c t i o n were i n C la s s 2 ( t a b le 1 3 ) . 1 0 th Day The d a ta i n t a b le 11 showed t h a t on th e 1 0 th day t h e m a jo r ity o f b ir d s showed a tum or r e a c t io n fo r l i n e s 6 , 7 , 9 , 1 0 , 1 1 , 1 4 , and 1 5 . The in c r e a s e d p e r c e n ta g e i n th e 1 , 2 , and 3 tumor c l a s s e s (ta b le 13) i s e v id e n c e o f t h e c o n tin u e d grow th o f th e t r a n s p la n t i n th e b ir d s i n a l l th e l i n e s . I t was n o t ic e d t h a t t h e p e r c e n ta g e o f b ir d s t h a t showed a grow th o f C la s s 2 was g r e a t e r in t h e s u r v iv o r s th a n in th e n o n -s u r v iv o r s in a l l l in e s excep t lin e 7• T h u s, s u s c e p t i b i l i t y or r e s i s t a n c e ca n n o t be d eterm in ed from t h e b r e a s t r e a c t i o n a lo n e d u r in g th e f i r s t t e n days a f t e r i n o c u l a t i o n . B r e a s t r e a c t i o n on d i f f e r e n t d a y s . G e n e r a tio n WKW 6t h Day The d a ta i n t a b l e 12 shows t h a t l e s s th a n 11 p e r c e n t o f t h e b ir d s i n l i n e s 4 , 6 , 7 , 9 , 1 1 , 1 4 , and 1 5 , r e s p e c t i v e l y , had b r e a s t r e a c t io n s on t h e 6 th day* 1 0 t h Day The r e a c t o r p e r c e n ta g e (b y t h e 1 0 th d a y ) in c r e a s e d i n ea ch l i n e . I t was 7 7 .6 , 8 1 .7 , 9 6 .4 , 9 5 . 2 , 6 2 . 5 , 9 1 .0 , and 8 5 .5 p e r c e n t i n l i n e s 4 , 6 , 7 , 9 , 1 1 , 1 4 , and 1 5 , r e s p e c t i v e l y . As a r e s u l t o f t h e c o n tin u e d grow th o f th e t r a n s p la n t t h e number o f r e a c t o r s in c r e a s e d in tumor c l a s s e s 1 , 2 , and 3 ( t a b l e l 4 ) • - 33 - The TABLE 12 The P e r c e n ta g e o f C hickens t h a t Showed V i s i b l e B r e a st Tumors i n t h e V a r io u s In b red L in e s o f G e n e r a tio n f,Ktt on t h e 6 th and 1 0 th Day a f t e r I n o c u l a t i o n . T o t a l N o. o f C h ick en s P e r c e n t Showing Tumors on 6 th Day P e r c e n t Showing Tumors on 1 0 th Day 4 139 1 .5 7 7 .6 6 328 1 0 .4 8 1 .7 7 190 9 .5 9 6 .4 9 186 1 1 .8 9 5 .2 11 64 4 .7 6 2 .5 14 122 9 .4 9 1 .0 15 117 0 .8 8 5 .5 L in e - 34 - TABLE 13 C la s s F req u en cy o f Breast; Tumor S iz e on "the 6 th and 1 0 th Day among T hose C h ick en s i n t h e V a r io u s In b red L in e s o f t h e " j ” G e n e r a tio n Which w ere A liv e on t h e lObh D ay, but D ie d P r i o r t o 28 Days a f t e r I n o c u l a t i o n . L in e Number of C h ick en s Tumor S iz e C la s s P e r c e n ta g e o f C h ick en s Show ing t h e S iz e o f Tumor on t h e — 6t h Day 1 0 th Day 6 69 0 1 2 3 8 1 .1 1 3 .0 5 .7 0 .0 1 1 .6 47 .8 3 1 .8 8 .6 7 37 0 1 2 3 9 1 .8 8 .1 0 .0 0 .0 13 .6 3 2 .4 3 7 .8 1 6 .2 9 41 0 1 2 3 4 6 3 .3 3 6 .6 0 .0 0 .0 0 .0 7 .3 2 1 .9 4 3 .9 2 4 .4 2 .4 10 93 0 1 2 3 6 6 .6 2 7 .9 5 .4 0 .0 4 .3 1 9 .3 5 0 .5 2 5 .8 11 81 0 1 2 3 8 2 .7 1 7 .2 0 .0 0 .0 2 4 .6 3 5 .8 3 5 .8 3 .7 14 40 0 1 2 3 6 0 .0 3 5 .5 5 .0 0 .0 1 0 .0 15 .0 4 2 .5 3 2 .5 15 172 0 1 2 3 8 1 .4 1 5 .6 1 -9 0 .0 9 .3 4 1 .2 41 .2 8 .2 - 35 - TABLE 14 C la s s F req u en cy o f B r e a s t Tumor S iz e on t h e 6 th and 1 0 th Day Among T hose C h ick en s i n t h e V a rio u s In b red L in e s o f th e G e n e r a tio n "Which w ere A liv e on t h e 1 0 th D ay, but D ied P r i o r t o 28 Days a f t e r I n o c u l a t i o n . L in e Number of C h ick en s Pe r c e n t age o f C h ick en s Show ing t h e S iz e o f 1Tumor on t h e — 1 0 th Day 6t h Day Tumor S i z e C la s s 4 32 0 1 2 3 1 0 0 ,0 0 .0 0 .0 0 .0 2 1 .8 4 3 .7 3 1 .2 3 .2 6 98 0 1 2 3 8 9 .3 1 0 .6 0 .0 0 .0 2 9 .6 5 2 .1 1 5 .9 2 .1 7 189 0 1 2 3 9 1 .0 8 .9 0 .0 0 .0 3 .2 5 9 .7 3 6 .5 0 .5 9 128 0 1 2 3 8 9 .0 1 0 .9 0 .0 0 .0 5 .4 5 8 .5 3 2 .1 3 .9 11 25 0 1 2 1 0 0 .0 0 .0 0 .0 3 6 .0 5 6 .0 8 .0 14 37 0 1 2 9 1 .3 8 .7 0 .0 14 .1 5 0 .8 3 .5 15 113 0 1 2 3 9 9 .1 .8 0 .0 0 .0 1 5 .0 6 9 .9 1 4 .2 0 .8 - 36 p e r c e n t a g e o f r e a c t o r s i n th e s u r v iv o r s o f ea ch c l a s s o f r e a c t i o n was n o t l e s s t h a n fo r th e c o r r e sp o n d in g c l a s s e s o f n o n - s u r v iv o r s . As i n g e n e r a t io n ”j " , t h e b r e a s t r e a c t i o n d id n o t d i s t i n g u i s h r e s is ta n t from s u s c e p t i b l e l i n e s a t e i t h e r 6 or 10 d a y s . B r e a s t r e a c t i o n o f s u r v iv o r s from th e 1 4 th t o 2 8 th d a y s . T a b le s 15 and 1 6 . A s tu d y o f t h e b r e a s t r e a c t io n i n t h e b ir d s t h a t s u r v iv e d u n t i l t h e end o f t h e ex p erim en t showed t h a t by t h e 1 4 th day a f t e r i n o c u l a t i o n , t h e b r e a s t r e a c t i o n i n ea ch l i n e had ta k e n a more d e f i n i t e p a t t e r n . I n some l i n e s t h e b ir d s showed tumor r e g r e s s i o n so t h a t t h e number o f n o n -r ea c to r s g r e a t ly in c r e a s e d . At t h e same tim e as a r e s u l t o f c o n tin u e d grow th o f tu m or i n some b ir d s t h e number o f r e a c t o r s in c r e a s e d i n c l a s s e s 3 , 4 , and 5 o f tum or s i z e . B reast r e a c tio n in d if fe r e n t l i n e s . L in e 6 : day. G e n e r a tio n MJ n . The r e g r e s s i o n i n t h e s i z e o f th e tumor began on t h e 1 4 th The number o f n o n - r e a c t o r s in c r e a s e d n o t i c e a b l y u n t i l on t h e 2 8 th day i t r e a c h e d 8 3 .4 p e r c e n t o f t h e t o t a l s u r v iv o r s . L in e 7 : O nly one b ir d d id not d ie from t h e tum or and i t grow th o f c l a s s 2 on t h e 1 4 th d a y . showed a T here was no e v id e n c e o f tum or i n t h e b r e a s t when t h e b ir d was exam ined on t h e 1 7 t h , 2 1 s t , and 2 8 th d a y . L in e 9 : G e n e r a lly r e g r e s s i o n i n th e s i z e o f th e tum or s t a r t e d by t h e 1 4 th day and by t h e 1 7 t h day f o r 20 p e r c e n t o f t h e b ir d s t h a t s u r v iv e d u n t i l t h e end o f t h e ex p erim en t a t w h ich tim e no tum ors w ere e v id e n t. The number o f b ir d s t h a t showed c o m p le te r e g r e s s i o n in c r e a s e d 7 3 .3 p e r c e n t on t h e f i n a l day o f t h e e x p e r im e n t. T here w ere no b ir d s sh ow in g a 3 , 4 , o r 5 c l a s s tum or growth, a t 28 d ays a f t e r i n o c u l a t i o n . - 37 - L in e 1 0 : Of t h e b ir d s t h a t rem ained on t h e f i n a l day o f th e e x p e r im e n t, 94*5 p e r c e n t d id n o t show any l o c a l i z e d tu m o r, h o w ev er, th e r e s t o f t h e b ir d s showed a tumor r e a c t i o n o f C la s s 5* W ith t h e number o f r e a c t o r s c o n t i n u a l l y d e c r e a s in g , t h e r e w ere no b ir d s t h a t showed C la s s 5 tu m o r s, e x c e p t on t h e f i n a l day o f t h e ex p erim en t* TJp t o th e 2 1 s t day none o f t h e b ir d s i n t h i s l i n e had a b r e a s t r e a c t i o n g r e a t e r th a n 4 d e g r e e s . L in e 1 1 : The d a ta i n t a b l e 12 f o r t h e 1 4 th day i n d i c a t e t h a t a l l o f t h e b ir d s had b r e a s t tum ors and 45 p e r c e n t o f t h e b ir d s were o f C la s s 4 tum or s i z e . On t h e 1 7 th day t h e p e r c e n ta g e o f b ir d s t h a t w ere i n 1 , 2 , 3 , and 4 c l a s s o f tum or s i z e was a p p r o x im a te ly e q u a l. A p p a r e n tly t h e b ir d s i n t h i s l i n e r e a c t e d s i m i l a r l y t o t h o s e o f l i n e 10 i n t h a t a c o m p a r a tiv e ly h ig h number o f b ir d s i n C la s s 5 tum or s i z e w ere p r e s e n t on t h e l a s t day o f th e e x p e r im e n t. L in e 1 4 : 1 4 th d a y . i h e s i z e o f t h e tum ors i n t h i s l i n e r e g r e s s e d a f t e r t h e On t h e 2 1 s t day 3*2 p e r c e n t o f t h e s u r v iv o r s were o f C la s s 5 tu m o r s, b u t t h e s e tu m ors r e g r e s s e d t o such an e x t e n t t h a t by t h e 2 8 th day none w ere la r g e r th a n a C la s s 2 r e a c t i o n . E ig h t y - s e v e n p e r c e n t showed no b r e a s t l e s i o n on t h e f i n a l d a y . L in e 15 : The tumor c o n tin u e d t o grow i n t h e b ir d s t h a t s u r v iv e d d u r in g t h e p e r io d o f e x p e r im e n t. A fte r t h e tumor had rea ch ed a grow th o f C la s s 4 i n some b i r d s , r e g r e s s i o n s t a r t e d and p r o g r e s s e d c o n t i n u a l l y . By t h e end o f t h e e x p e rim en t 7 1 .4 p e r c e n t o f t h e s u r v iv o r s d id n o t show tumor g ro w th , w h ile t h e r e s t showed C la s s 1 turner g r o w th . - 38 - B re a st r e a c tio n in d if f e r e n t lin e s * L in e 4 ; Wo b ir d s i n t h i s l i n e d e v e lo p e d a C la s s 5 grow th d u r in g t h i s e x p e r im e n t. o f C la s s 4 . G e n e r a tio n "K" On th e 17 th d a y 8 .4 p e r c e n t o f t h e b ir d s had a tumor T h is was t h e h ig h e s t p e r c e n ta g e o f r e a c t o r s i n l i n e 4 t h a t r e a c h e d t h i s c l a s s d u r in g t h e p e r io d o f t h e e x p e r im e n t. S t a r t in g from t h e 1 4 th d a y t h e number i n t h e upper c l a s s e s o f tum or r e a c t io n d e c r e a s e d c o n t i n u a l l y and by t h e 2 8 th day 9 5 .4 p e r c e n t o f t h e s u r v iv o r s d id n o t show any g r o w th . A v e r y s m a ll p e r c e n ta g e o f t h e b ir d s showed a l o c a l r e a c t i o n o f 1 , 2 , and 3 on th e f i n a l day o f t h e e x p e r im e n t. L in e 6 : On th e 1 4 th d ay th e number o f 0 C la s s i n t h i s l i n e in c r e a s e d i n c o m p a r iso n w ith t h e same p e r c e n ta g e s bn t h e 1 0 th d a y . b ir d s w ere i n C la s s 1 . Won© o f t h e The number o f b ir d s i n C la s s 2 d e c r e a s e d and t h e number o f b ir d s i n C la s s e s 3 , 4 , and 5 i n c r e a s e d . On t h e 1 7 t h , 2 1 s t and 2 8 th day t h e number o f b ir d s i n ea ch c l a s s o f grow th d e c r e a s e d c o n tin u a lly * T h is shows t h a t t h e tum ors i n t h e b ir d s o f t h i s l i n e r e a c h e d t h e p o in t o f maximum grow th by th e 1 4 th day* L in e 7 : s u r v iv e d . S im ila r t o t h e same l i n e in g e n e r a t io n "J” , one b ir d T h is b ir d showed a tum or grow th o f C la s s 1 on t h e 1 4 th d a y . The tum or had r e g r e s s e d c o m p le te ly when th e b ir d was exam ined on t h e 1 7 th day • L in e 9 : The number o f b ir d s t h a t d id not show tum ors began t o in c r e a s e by t h e 1 4 th d a y . t o r e g r e s s by t h e 1 4 th d a y . T h is shows t h a t m ost o f t h e tum ors s t a r t e d H owever, i n a few b ir d s t h e tumor c o n tin u e d t o grow . - 39 - TABLE 15 C la s s F req u en cy o f B rea st Tumor S iz e among t h o s e C h ick en s i n th e V a r io u s In b red L in e s o f t h e 11J ” G e n e r a tio n w h ich S u r v iv e d u n t i l 28 Days a f t e r I n o c u l a t i o n - P e r c e n ta g e o f C h ick en s Show ing th e S iz e o f Tumor on th e -* 1 4 th 1 7 th 21st 2 8 th 6t h 1 0 th Day Day Day Day Day Day No • o f C h ick en s Tumor S iz e C la s s 6 217 0 1 2 3 4 5 6 7 .7 2 9 .0 3 .2 0 .0 0 .0 0 .0 9 .6 4 3 .3 3 6 .8 1 0 .2 0 .0 0 .0 1 0 .5 1 2 .9 3 5 .4 3 0 .5 1 0 .6 0 .0 3 7 .5 1 5 .6 1 7 .9 1 5 .2 1 3 .3 0 .4 6 1 .7 1 1 .0 1 2 .5 6 .5 7 .8 0 .4 8 3 .4 1 2 .4 2 .7 0 .5 0 .4 0 .4 7 1 0 1 2 1 0 0 .0 0 .0 0 .0 0 .0 1 0 0 .0 0 .0 0 .0 0 .0 1 0 0 .0 1 0 0 .0 0 .0 0 .0 1 0 0 .0 0 .0 0 .0 1 0 0 .0 0 .0 0 .0 9 15 0 1 2 3 4 6 6 .6 2 0 .0 1 3 .3 0 .0 0 .0 6 .7 1 3 .3 66 .6 1 3 .3 0 .0 0 .0 1 3 .3 2 6 .6 2 0 .0 1 4 .0 2 0 .0 2 0 .0 3 3 .3 2 0 .0 6 .6 6 0 .0 2 0 .0 1 3 .3 6 .6 0 .0 7 3 .3 2 0 .0 6 .6 0 .0 0 .0 10 37 0 1 2 3 4 5 6 4 .8 2 9 .7 5 .4 0 .0 0 .0 0 .0 5 .4 1 8 .9 4 8 .6 2 4 .3 2 .7 0 .0 0 .0 5 .4 2 1 .6 5 4 .0 1 8 .9 0 .0 1 6 .2 32 .4 2 1 .6 1 6 .2 13 .5 0 .0 5 9 .4 2 4 .3 5 .4 8 .1 2 .7 0 .0 9 4 .5 0 .0 0 .0 0 .0 0 .0 5 .4 11 40 0 1 2 3 4 5 7 7 .5 2 2 .6 0 .0 0 .0 0 .0 0 .0 15 .0 3 5 .0 4 5 .0 5 .0 0 .0 0 .0 0 .0 2 .5 1 2 .5 2 0 .0 45 .0 2 0 .0 1 2 .5 2 2 .5 1 7 .5 2 2 .5 2 2 .5 2 .5 4 0 .0 2 0 .0 7 .5 1 0 .0 1 7 .5 5 .0 7 0 .0 5 .0 1 2 .5 2 .5 0 .0 1 0 .2 14 31 0 1 2 3 4 5 6 7 .7 2 9 .0 3 .2 0 .0 0 .0 0 .0 3 .3 1 9 .3 5 8 .0 1 9 .3 0 .0 0 .0 0 .0 6 .5 1 9 .3 4 8 .3 2 5 .8 0 .0 9 .6 1 9 .5 2 2 .5 2 2 .5 2 5 .8 0 .0 3 8 .7 2 2 .5 1 9 .3 9 .7 6 .5 3 .2 8 7 .0 9 .7 3 .2 0 .0 0 .0 0 .0 7 0 1 2 3 4 5 6 .1 2 8 .5 1 4 .2 0 .0 0 .0 0 .0 1 4 .3 4 2 .8 4 2 .8 0 .0 0 .0 0 .0 4 2 .9 2 8 .5 2 8 .5 2 8 .5 0 .0 1 4 .4 2 8 .5 2 8 .5 2 8 .5 1 4 .3 4 2 .8 1 4 .3 0 .0 7 1 .4 2 8 .5 0 .0 0 .0 0 .0 L in e 15 - 40 - TABLE 16 C la s s F req u en cy o f B re a st Tumor S iz e among t h o s e C h ick en s i n t h e V a r io u s In b red L in e s o f t h e "KM G e n e r a tio n w h ich S u r v iv e d u n t i l 28 D ays a f t e r I n o c u l a t i o n . P e r c e n ta g e o f C h ick en s Showing th e S iz e o f Tumor on t h e — 2 8 th 21st 6t h 1 4 th 1 7 th 10t h Day Day Day DayDay Day Wo • o f C h ick en s Tumor S iz e C la s s 4 107 0 1 2 3 4 9 8 .1 1 .8 0 .0 0 .0 0 .0 2 2 .4 5 1 .4 2 5 .2 0 .9 0 .0 2 4 .3 2 3 .4 3 7 .3 14 .0 0 .9 4 7 .7 1 5 .8 1 9 .6 8 .4 8 .4 7 8 .5 1 2 .1 0 .9 5 .6 2 .8 9 5 .4 1 .8 1 .8 0 .9 0 .0 6 230 0 1 2 3 4 5 8 9 .7 9 .8 0 .4 0 .0 0 .0 0 .0 1 3 .7 4 7 .8 3 6 .7 1 .7 0 .0 0 .0 2 0 .0 0 .0 2 0 .5 4 1 .5 1 7 .5 0 .4 4 7 .8 0 .0 1 9 .2 1 8 .8 1 1 .5 0 .2 5 6 6 .7 1 6 .7 1 0 .7 2 .9 2 .9 0 .0 9 1 .4 5 .5 1 .7 0 .0 1 .3 0 .0 7 1 0 1 0 .0 1 0 0 .0 1 0 0 .0 0 .0 0 .0 1 0 0 .0 1 0 0 .0 0 .0 1 0 0 .0 0 .0 1 0 0 .0 0 .0 9 58 0 1 2 3 4 8 6 .2 1 3 .7 0 .0 0 .0 0 .0 3 .4 4 8 .4 4 4 .8 4 .3 0 .0 1 2 .0 1 7 .4 5 5 .1 1 3 .7 1 .7 3 4 .4 2 0 .6 2 9 .3 1 2 .0 3 .4 6 8 .9 1 3 .8 1 2 .0 5 .2 0 .0 9 6 .5 1 .7 0 .0 0 .0 1 .7 11 39 0 1 2 3 4 9 2 .3 7 .6 0 .0 0 .0 0 .0 3 8 .5 4 3 .5 1 7 .9 0 .0 0 .0 1 5 .3 2 5 .6 4 3 .5 1 0 .3 5 .2 3 3 .3 2 .5 3 0 .8 2 3 .0 1 0 .3 5 1 .3 1 0 .3 1 7 .9 7 .6 1 2 .8 8 2 .0 7 .8 2 .5 2 .5 5 .1 14 75 0 1 2 3 4 5 9 0 .7 9 .2 0 .0 0 .0 0 .0 0 .0 4 .6 6 0 .0 3 5 .4 0 .0 0 .0 0 .0 1 6 .9 12 .4 5 3 .8 1 5 .4 1 .4 0 .0 4 0 .0 1 2 .4 1 6 .9 2 3 .0 7 .6 0 .0 7 5 .3 4 .7 1 0 .7 4 .7 3 .0 1 .5 8 9 .2 9 .2 0 .0 0 .0 1 .6 0 .0 15 4 0 1 2 1 0 0 .0 0 .0 0 .0 0 .0 75 .0 2 5 .0 0 .0 2 5 .0 7 5 .0 1 0 0 .0 0 .0 0 .0 1 0 0 .0 0 .0 0 .0 1 0 0 .0 0 .0 0 .0 L in e - 41 - L in e 1 1 : I n t h i s l i n e some tumor r e g r e s s i o n s o ccu rred b y t h e 1 7 th d a y . The b ir d s showed t h e same grow th tr e n d as t h o s e of* l i n e 9 e x c e p t t h a t on t h e 2 8 th day r e a c t o r s o ccu rred i n a l l c l a s s e s e x c e p t F iv e and o n e - t e n t h p e r c e n t o f t h e b ir d s w ere i n C la s s 4 . 5. T h is i s a f a i r l y h ig h p e r c e n ta g e when compared t o th e p e r c e n ta g e i n t h i s c l a s s i n t h e o th e r l i n e s . L in e 1 4 : The d a ta show t h a t t h e p e r c e n ta g e i n a l l c l a s s e s o f grow th, e x c e p t 5 , d e c r e a s e d a f t e r i t r ea c h e d i t s maximum on th e 1 7 th day. On t h e l a s t day o f t h e ex p erim en t one b ir d out o f 75 s u r v iv o r s showed a C la s s 4 tum or s i z e . T h is was t h e o n ly b ir d w ith a la r g e r t h a n C la s s 1 tu m o r . L in e 1 5 : O nly 3*4 p e r c e n t o f t h e s e b ir d s (4 out o f 1 1 7 ) l i v e d t h e end o f t h e ex p erim en t • to On t h e 1 0 th and 1 4 th d ay 25 p e r c e n t and 75 p e r c e n t o f t h e s e b ir d s showed c l a s s e s 1 and 2 b r e a s t r e a c t i o n s . On t h e 1 7 th d ay t h e tum ors had c o m p le te ly r e g r e s s e d i n s i z e . I n g e n e r a l , i t was n o t ic e d t h a t t h e r e was a p e r io d i n w h ich t h e t r a n s p la n t s t a r t e d and c o n tin u e d grow ing fo llo w e d by a r e g r e s s i o n i n s iz e . T here w ere a l s o n o t ic e a b l e v a r i a t i o n s b etw een t h e d i f f e r e n t l i n e s in r e g a r d t o tumor grow th and r e g r e s s i o n . B ir d s i n some l i n e s had an e a r l i e r s t a r t i n grow th o f tumor t h a n o t h e r s . I t was a l s o o b serv ed t h a t t h e d i s t r i b u t i o n o f b ir d s i n C la s s 5 f lu c t u a t e d more i n some l i n e s th a n in o th e r s . The n a tu r e o f t h e tumor i n th e b r e a s t m u scle on t h e 2 8 th day was d i f f e r e n t th a n t h e tu m ors o f s i m ila r s i z e i n t h e b ir d s t h a t d ie d e a r l i e r . Tumors showed more n e c r o t i c t i s s u e w ith th e p r o g r e s s o f t i m e . T hose o f C la s s 1 and C la s s 2 on t h e 2 8 th day w ere m o s tly a c a p s u le c o n t a in in g n e c r o tic t i s s u e . - 42 - No g r o s s a l t e r a t i o n was found a t t h e s i t e o f in o c u la tio n in th e b ir d s m w h ich t h e t r a n s p la n t c o m p le te ly r e g r e s s e d * A v a r ia b le amount o f n e c r o s i s was commonly found i n t h e tum ors o f b ir d s t h a t d ie d a t l a t e r a g e s* I n some in s t a n c e s la r g e tum ors w ere a lm o st e n t i r e l y in v o lv e d b y n e c r o s i s and i n su c h c a s e s t h e b ir d s were found t o be d u l l , l i s t l e s s , and t h e y r e f u s e d t o e a t • METASTATIC INVOLVEMENT OF THE INTERNAL ORGANS A s id e from l o c a l grow th o f t h e im p la n t i n t h e b r e a s t m u s c le s , a d i f f u s e ty p e o f m e t a s t a s is was found i n t h e v i s c e r a * The o rgan s most f r e q u e n t l y in v o lv e d w ere t h e l i v e r , k id n e y , s p le e n , h e a r t , p a n c r e a s , p r o v e n t r i c u l u s , and g o n a d s. The o rg a n s a re ranked i n t h e i r o rd er o f fr e q u e n c y o f in v o lv e m e n t. L iv e r : I n t h e l i v e r t h e m e t a s t a s is m a n ife s te d i t s e l f i n tw o t y p e s , a " d iffu se** (ty p e A) and a "nodular" (ty p e B ) . I n th e form er t y p e , w h ich w as t h e mo ne common, t h e l i v e r was e n la r g e d t o v a r io u s d e g r e e s . I n t h e b ir d s t h a t d ie d a sh o r t tim e a f t e r i n o c u l a t i o n , t h e l i v e r showed c o n s id e r a b le c o n g e s t io n and i n some i n s t a n c e s i t rea ch ed t w ic e t h e norm al s iz e . B ir d s t h a t d ie d d u r in g t h e l a t e r p e r io d o f t h e ex p erim en t had a d i f f u s e or f o c a l in v o lv e m e n t o f t h e l i v e r w ith l i t t l e in c r e a s e i n t h e s i z e o f t h i s o rgan • K id n e y s: T h e se organ s w ere p r im a r ily a f f e c t e d by d i f f u s e g r e y is h e n la r g em en t o f some o f t h e t h r e e l o b e s . p u f f y and e n la r g e d . I n some c a s e s t h e y w ere n o t ic e a b l y I n g e n e r a l t h e in v o lv e m e n t o f t h e k id n e y s p a r a l l e l e d th e s e v e r ity o f liv e r ch an ges. S p le e n : th is I n b i r d s t h a t d ie d w it h in t h e f i r s t week a f t e r i n o c u l a t i o n o r g a n was e n la r g e d . c o n t a in tu m o r n o d u le s . I t had a u n ifo rm c h e r r y -r e d c o lo r and did n o t I n t h e b i r d s t h a t d ie d l a t e r , t h i s o rg a n was - 43 - u s u a l l y l e s s c o n g e s t e d , g r a y ish -b r o w n i n c o l o r , s o f t i n t e x t u r e , and showed some d e g r e e o f a tr o p h y . H e a r t: I n t h e b ir d s t h a t d ie d e a r l y , a d i f f u s e m y o c a r d itis o c cu rr e d and t h e organ w as s l i g h t l y e n la r g e d . The b ir d s t h a t d ie d i n t h e l a t e r p e r io d o f t h e ex p e rim en t showed prom inent m y o c a r d ia l tu m ors w h ich w ere som etim es accom panied by p e r i c a r d i t i s • T h ese tum ors u s u a l l y w ere f o c a l and o f t e n showed c o a l e s c e n c e . P an creas : T h is organ was o f t e n a f f e c t e d w ith f o c a l tu m o r s. The m e t a s t a t i c f o c i w ere u s u a ll y c ir c u m sc r ib e d and e a s i l y d e t e c t e d upon g r o s s e x a m in a t io n . In su ch c a s e s t h e p a n crea s was th ic k e n e d and f ir m . Some in v o lv e m e n ts o f t h e p a n cr e a s may have b een o v e r lo o k e d b e c a u se t h e c o lo r o f t h i s organ i s s im ila r t o t h e c o lo r o f t h e tumor t i s s u e . In t h e b ir d s t h a t d ie d d u r in g l a t e r s t a g e s t h e tum ors i n th e p a n cr e a s w ere q u ite s m a ll i n s i z e and gave t h e im p r e s s io n t h a t t h e y had undergone r e g r e s s io n * P r o v e n tr ic u lu s : D if f u s e i n f i l t r a t i o n o f th e n e o p l a s t i c lym phoid t i s s u e w as a c h a r a c t e r i s t i c f in d in g i n t h e w a ll o f t h e p ro v en t r i c u l u s , h o w e v e r, t h i s c o n d i t i o n was n o t n o t ic e d e x c e p t i n b ir d s t h a t d ie d i n th e l a t e r p e r io d o f t h e e x p e r im e n t. The w a ll o f t h e p ro v en t r i c u l u s was g r e a t l y th ic k e n e d and n e c r o t i c a t t i m e s . G onads: The imm ature ovary showed an in c r e a s e i n s i z e . i n s t a n c e s t h e y w ere t h r e e tim e s t h e i r normal s i z e . w ere som etim es e n la r g e d and were v e r y f ir m . I n some The m ale g la n d s The in v o lv em en t o f t h e s e organ s u s u a l l y o c c u r r e d i n b ir d s t h a t d ie d i n t h e l a t e r s t a g e s o f t h e ex p e rim en t • P e r ito n e u m : str u c tu r e . I n v e r y few c a s e s was t h e d is e a s e m a n ife s t e d i n t h i s The tum ors t h a t e x i s t e d w ere d i f f u s e i n n a t u r e . - 44 - Lungs^: T here w ere no s i g n i f i c a n t ch an ges i n t h e lu n g s • In some c a s e s t h e c a r c a s s was m arkedly e m a c ia te d . L iv e r m e t a s t a s i s : (T a b le s 17 and 1 8 ) I n g e n e r a t io n "J" th e v a r io u s l i n e s o f b ir d s ca n be ranked a s f o l l o w s i n o rd er o f t h e i r s u s c e p t i b i l i t y t o l i v e r in v o lv em en t : 7 , 1 5 , 1 0 , 1 1 , 9 , 1 4 , and 6 . The l i n e s f e l l i n t o t h r e e g r o u p s . The most s u s c e p t i b l e group was composed o f l i n e s 7 and 1 5 , w hich had a p p r o x im a te ly t h e same p e r c e n ta g e of' in v o lv em en t (9 6 p e r c e n t ) . The secon d s u s c e p t i b l e group in c lu d e d l i n e s 1 0 , 1 1 , and 9 w it h a p p r o x im a te ly 70 p e r c e n t i n v o l v e ­ m e n t. The t h i r d group c o n ta in e d r e s i s t a n t l i n e 14 w it h a p p r o x im a te ly 57 p e r c e n t and l i n e 6 w ith a p p r o x im a te ly 23 p e r c e n t . I n g e n e r a l, t y p e "B” l i v e r involvem ent: was found i n t h o s e b ir d s t h a t te n d e d tow ard r e s i s t a n c e t o th e tum or • I n g e n e r a t io n "K,f l i n e s 7 and 15 were a g a in th e m ost s u s c e p t i b l e l i n e s h a v in g an in c id e n c e o f l i v e r in v o lv em en t o f 99*4 p e r c e n t and 9 6 .5 p e r c e n t . L in e 9 had t h e same in c id e n c e i n b o th g e n e r a t io n s . L in e s 11 and 14 showed l e s s in v o lv em en t o f th e l i v e r i n g e n e r a t io n "Kw th a n i n g e n e r a t io n " j " . L in e 4 was r e p r e s e n te d i n th e "K” g e n e r a t io n o n ly and l i n e 10 was a b s e n t from t h i s g e n e r a t io n . The l i v e r in v o lv e m e n t in l i n e 6 was s l i g h t l y but n o t s i g n i f i c a n t l y h ig h e r (28 p e r c e n t ) th a n i n g e n e r a t io n "J" . T h u s, In t h e "K" g e n e r a t io n t h e l i n e s ranked f o r l i v e r in v o lv e m e n t a s f o l l o w s : 7 , 1 5 , 9 , 1 4 , 1 1 , 6 , and 4 . M e t a s t a t ic in c id e n c e o f th e i n t e r n a l o r g a n s : The p e r c e n ta g e o f b ir d s i n v a r io u s l i n e s t h a t had tum or in v o lv em en t o f th e k id n e y , s p le e n h e a r t , p a n c r e a s , p r o v e n t r i c u l u s , g o n a d s, and p erito n eu m i s l i s t e d i n t a b l e 19 fo r g e n e r a t io n and t a b l e 20 fo r g e n e r a t io n nKw. - 45 - TABLE 17 T ypes oT L iv e r Involvem ent; i n G e n e r a tio n UJ H L in e 6 Type P e r c e n ta g e o f B ir d s L iv in g 28 Bays P e r c e n ta g e o f B ird s L iv in g L e ss th a n 28 Days T o ta l P e r c e n ta g e 5 .7 8 8 .5 5 .7 7 6 .5 2 2 .0 1 .4 9 7 .2 2 .7 2 .6 9 4 .7 2 .6 - 4 .8 9 0 .3 4 .8 3 0 .4 6 6 .0 3 .5 0 A B 9 4 .5 5 .4 - 2 .1 9 5 .7 2 .1 28 .4 7 0 .0 1 .5 11 0 A B 8 7 .5 1 2 .5 - 2 .4 9 3 .8 3 .7 3 0 .5 6 6 .9 2 .5 14 0 A B 9 6 .7 3 .2 0 A B 1 0 0 .0 0 A B 9 9 .0 0 .9 0 A B 1 0 0 .0 0 A B 1 0 0 .0 10 7 9 15 0 * - - - - — - No r e a c t i o n - 46 - _ 1 0 0 .0 — - 9 8 .8 1 .1 4 2 .2 5 7 .7 3 .9 9 4 .9 1 .1 TABLE 18 T ypes o f L iv e r In v o lv em en t i n G e n e r a tio n UKI! L in e 6 7 9 4 11 14 15 Type P e r c e n ta g e o f B ird s L iv in g 28 Days 0 A B 9 9 .1 0 .8 0 A B 1 0 0 .0 0 A B 1 0 0 .0 0 A B 9 7 .1 1 .9 0 .9 0 A B 92 .3 7 .6 - 0 A B 9 6 .9 3 .0 0 A B 1 0 0 .0 - - - - - - P e r c e n ta g e o f B ir d s L iv in g L e s s th a n 28 Days 4 .3 8 6 .2 9 .5 7 1 .9 2 5 .3 2 .7 9 7 .8 2 .1 0 .5 9 7 .3 2 .1 0 .7 9 6 .8 2 .4 3 1 .7 6 6 .6 1 .6 9 0 .6 9 .3 7 4 .8 2 2 .3 2 .8 9 2 .0 8 .0 5 6 .2 4 0 .6 3 .1 _ — 1 0 0 .0 - - — 1 0 0 .0 ■ 0 = No r e a c t i o n - 47 - T o ta l P e r c e n ta g e 5 1 .6 4 8 .3 3 .4 9 6 .5 I n g e n e r a l i n g e n e r a tio n s MJ n and MKn t h e r e m s c l o s e agreem ent on t h e p e r c e n ta g e o f b ir d s w it h in th e v a r io u s l i n e s w h ich had i n v o l v e ­ ment o f t h e k id n e y , s p le e n , and h e a r t* L ik e w is e , t h e s e p e r c e n t a g e s w ere i n c l o s e agreem ent w ith t h e p e r c e n ta g e s t h a t were o b se rv e d fo r l i v e r in v o lv e m e n t . I t was o b se rv e d t h a t t h e p a n c r e a s , p r o v e n t r ic u i u s , g o n a d s, and p e r ito n e u m were l e s s f r e q u e n t ly in v o lv e d th a n t h e o th e r organ s and t h e p e r c e n t a g e o f b ir d s t h a t had tum ors o f t h e s e organs was e x tr e m e ly v a r i a b l e and no r e l a t i o n t o s u s c e p t i b i l i t y o r r e s i s t a n c e c o u ld be d e te c te d • T h ese org a n s (p a n c r e a s , p r o v e n t r ic u iu s , and g o n a d s) were u s u a l l y more s e v e r e l y in v o lv e d i n t h e b ir d s t h a t succumbed t o th e tumor but l i v e d lo n g e r b e fo r e d y in g . I n g e n e r a l t h e p e r c e n ta g e o f b ir d s t h a t showed in v o lv em en t o f t h e s e o rg a n s was h ig h e r i n g e n e r a t io n - 48 - th a n i n g e n e r a t io n MKTt. DISCUSSION I n t h e s e e x p e r im e n ts , t h e r e s u l t s o f in tr a m u s c u la r t r a n s p l a n t a ­ t i o n w ith non—f i l t e r e d tum or s t r a i n RPL 1 6 , i n d i c a t e a s i m i l a r i t y i n t h e r e a c t i o n o f t h e v a r io u s l i n e s t o t h a t o f t h e n a t u r a l l y o c c u r r in g d is e a s e . The d a t a i n t a b l e 5 on "the r e s p o n s e o f t h e c h ic k e n s i n t h e s e v e r a l l i n e s t o tum or t r a n s p l a n t s o f RPL 16 show t h a t t h e r e i s a g r e a t d i f f e r e n c e i n r e s i s t a n c e betw een l i n e s . The l i n e s w hich had b e e n s e l e c t e d u n d e r n a t u r a l ex p o su re f o r r e s i s t a n c e o r s u s c e p t i b i l i t y t o ly m p h o m a to sis , i n g e n e r a l , fo llo w e d t h e same tr e n d i n r e g a r d t o r e s i s t a n c e o r s u s c e p t i b i l i t y to t h e e x p e r im e n ta lly in d u c e d tu m o rs . L in e 6 w hich was r e l a t i v e l y r e s i s t a n t t o t h e n a t u r a l i n f e c t i o n was l i k e w i s e q u i t e r e s i s t a n t to i n o c u l a t i o n w ith tum or t r a n s p l a n t s . H ow ever, l i n e 10 w hich h a d shown c o n s id e r a b le r e s i s t a n c e to n a t u r a l l y o c c u r r i n g ly m p h o m a to sis, t*as q u i t e s u s c e p t i b l e t o tum or t r a n s p l a n t s . L in e s 7 , 9» a n ^ 15 w ere t h e m ost s u s c e p t i b l e l i n e s . D ata c o l l e c t e d o n n a t u r a l i n f e c t i o n o v e r a p e r io d o f 600 days ( t a b l e 21) f o r s i b s o f t h e i n o c u l a t e d b i r d s i n g e n e r a t i o n “J ” i n d i c a t e d t h a t t h e s e l i n e s r e a c t e d s i m i l a r l y t o b o th ty p e s o f i n f e c t i o n . The same co m p ariso n f o r s i b s i n g e n e r a t i o n UKH was n o t a v a i l a b l e a t 600 d ay s o f a g e . The s ib s o f b i r d s s e l e c t e d f o r s u s c e p t i b i l i t y to t h e n a t u r a l d is e a s e f e l l i n th e same c a te g o r y w hen i n o c u l a t e d w ith t h e t r a n s p l a n t . However, t h e l i n e s d id n o t f o llo w e x a c t l y t h e same o r d e r i n t h e two c a s e s . W ith t h e e x c e p tio n o f l i n e 6 , t h e d e a th r a t e o f t h e i n o c u l a t e d b i r d s was a b o u t t w ic e a s h ig h a s t h e d e a th r a t e o f t h e i r r e s p e c t i v e s i b s ex p o sed t o t h e n a t u r a l l y —o c c u r r in g d i s e a s e . I t h a s b een r e p o r t e d by J o h n s o n ( 1 9 5 7 ), J u n g h e r r (1 9 5 7 , Lee and W ilcke (1 9 ^ 1 ) and DeOme (1 9 ^ 5 ) - 2*9 - oj rH _ d LfN 0 - ON OJ »-3 • O ON KN ONxO CVJ C" u a © 03 M o aJ C Wo 03 • rH i—1 xO ON OJ a o rH o © id xO KN OJ UN CVJ U N -d CVI i 1 xO UN xO c- cvi -d KN • rH UN KN UN rH O OJ t —xO KNX0 O On ONx£> • CO ON KN O - ON KN rH KN fx- CO * xO CO UN C— kn UN OJ KNxO CVI o ON O - 00x0 UN O CVJ CO - d CO oj o CVJ CO ON KN CVJ rH KN _ d kn CVJ UN K N -d UN KN xO OJ KN ON cvi r-~ CVJ c^CVJ xo O -xO KNxO xO ON rH OJ ON KN rH KN tx - O • O KN ON i— 1 o ON 0 - o ON O KN KN O rH KN CD CO xO rH rH £xxO 0J xO ON rH CVJ rOv i—t • ON °Cx- ON rH -d . * 0 0 rH KN KN -d Cx- ON rH O UN rH C— OJ KN o- cA d 00 CVI o xO ON rH xO _d cu rH CN- OJ ON rH CVI • CO O N -d KN UN NN . ON 00 UN rH x 0 rH CVI ON • o x -d KN UN rH O x6 o rH - d XO KN OJ NN rH CVJ UN K N -d C~- IS’^ Cx- CO rH KN KN UN KN r - cA KN ON _d oj UN IN­ ~d o o OJ -d UN UN 00 UN • KN CD CO xO ON O Ex- O rH rH O 00 C— ON f- o UN f — f — UN LfN OJ KN ON xo o xO U N _d xQ O OJ CVI UN KN Pi rH UN rH rH OJ OJ ON ft CO KN C— UN xO UN [X- ON tx- UN KN C*CVI O OJ OJ rH xQ OJ fx• KN Q U N _d rH 0J >0 S CD s i fx-CO OJ _ d CO i rH _ d i—1 O • xQ rH c o r~_ • O •H 03 x0 ON KN OJ OJ a -d rH i—1 • KN LfN CO 0J ON UNCO KN OJ O rH r—t UN ON x£) O O KN O UN OJ <-H ON UN 00 xO CD KN OJ O - Q KN _ d rH cu Kidney ON sa? o fH o O ■— I o x0 rH KN K N -d of birds o C• rH rH ON CxrH number Perit • CVJ CVI UN rH indicates xO OJ O KN rH & of figures CD KN •P ©I i— rO •H CO •H > In each line fir s t set percentage of b ir d s . xQ CO CVI 121 Total . o O O * ON rH KN i-H OJ cR KN 00 • i-H O t — CVI G onad « CVI 00 ■ rH [— CVJ ON KN CVJ c— cu rH i—1 OJ ^ CVJ CO rH x£> KN O ON KN pH CVI rH H o xo o 00 o rH i—1 KN CVI x o rH o xO o CVJ CO rH rH CVI o xo CVJ rH O O • • o o KN O CO CM O n rH LA ON OJ OJ OJ - 4 CO UN CVI O rH - 4 CVJ KN LA CVI KN OJ LfN • xO rH KN f rH x 0 CO cvj o t- O rH O rH rH i—i LA CO 0 - LfN O'- ON 1-H o CVI o cvj o 1—1 1—1 CVI CO KN A - ON rH o o -4 KN CO xO O n t— 0 \-4 o tN- o rH O i-H rH O 7 ? ^ KN IN - ON 00 O- xO ON CVJ CVI O s rH _ 4 UN i—i O n KN CO k n r — on r*H On O xO ON i—1 C"• KN KN KN CVJ CVJ ON i—I xO xQ CO CVJ CO CO 00 ON I—1 KNxO CVI xO 1—1 rH set O O ACVJ rH 00 CN xO xO O CO p—I A - CM ON CO rH CO LA CO 00 rH CVI CVJ CK rH KN CVI rH o CVI XO rH rH XO KN i—1 rH and second C— O drH © H 04 o CVJ O at -P O J rH vO d O xQ r—f CN ON LfN LfN OO CVJ 0 0 00 00 ON rH f - £ © © f4 # KN KN KN CVJ i-H Oh CO O CVJ O K N _4 o P4 CVI KN KN CVI LfN • ON C— xO CO CVJ CO 0 0 0 0 ON ON O © £ -H .4 C ^xO O t— i—l -4 rH xO rH C*"~ S r i 9 -i 4 xO LA CN O 00 rH rH rH rH ACM L A rH ON rH xO • CM KN rH xQ LA L A -4 ON ON O KNxO CM CVJ o KN O xO KN A- x£> O CVI -4" ON CD LA _4 CM UN rH ON xO CM KN rH LA _4 xO rH o xO LA oj LA ON CVI KN -4 xO LA xO KN OO 00 C—- d KN LA O KN KN CVI rH CVJ KN xO KN 00 ON KN L A xO LA r— ON rH rH 3 -4 i—1 KN CVJ x 0 rH rH LA O • CVI rH c J^ - rH 00 • KN KN x 0 KN -4 number CO O O rH ON rH ind icates • r t rH 00 figures W O 00 i-H of S (U fl «© o u © fQ o (4 •H «H O © uo o • O i-H ON rH rH 00 0 \ KN ON rH H e a rt Eh O K id n e y 1 PQ £ o u Ph & <>H 09 u £ •H vO LA o o rH of birds P4 -P at PQ o -P © m > o 525 •rl -P © i—I £ w •rH > LA rH P4 In each line fir s t set percentage of b ir d s . £j o •H 0 cu -4 rH O of O U © £ O r■—i i-H in d ic a te s O O On rH figures 04 00 CVI KN 122 rH 186 T o ta l £ u © CQ • o S23 TABLE 21 C om parison o f P e r c e n ta g e M o r t a lit y fo r N a t u r a lly O ccu rrin g and T ran s p la n t ed Lymphomat o s i s . N a t u r a lly O ccu rrin g Lym phom atosis G e n e ra tio n s - i t j i r U J II L in e s — T r a n sp la n te d Tumor G e n e ra tio n s K 2 9 .8 1 7 .8 2 4 .1 10 2 4 .7 1 2 .1 7 1 .5 - 11 1 9 .8 - 6 6 .9 3 9 .0 14 2 4 .4 2 0 .9 5 6 .3 4 6 .7 4 2 6 .6 - 2 3 .0 7 4 2 .9 3 3 .1 9 7 .4 9 9 .5 4 0 .0 7 3 .2 6 8 .8 5 1 .7 96 .6 9 6 .5 9 15 in. 9 .3 to 6 5 6 .1 - - 52 t h a t t h e in c id e n c e o f tumor f o llo w in g in o c u l a t i o n was more th a n i n t h e ca se o f n a tu r a l in fe c tio n * I t w i l l be o b serv ed i n t a b l e 22 t h a t from y e a r t o y e a r c o n s id e r a b le v a r i a t i o n i n m o r t a li t y from n a t u r a lly o c c u r r in g ly m p h o m a to sis was p r e s e n t i n a l l l i n e s * I n g e n e r a l t h e r e was l e s s f l u c t u a t i o n i n b o th g e n e r a t io n s 11J M and 11Kw fo r t h e same l i n e s when in o c u l a t e d w it h tum or t r a n s p la n t s * The d i f f e r e n c e i n t h e d i s e a s e in c id e n c e from y e a r t o y e a r under t h e n a t u r a l c o n d it io n s may i n p a r t be a t t r ib u t e d t o t h e d i f f e r e n c e i n th e i n f e c t i v e d ose w it h in and b etw een y e a r s . A pp arent d i f f e r e n c e s b etw een t h e l i n e s s u g g e s t a d e g r ee o f g e n e t ic i d e n t i t y w i t h i n e a ch l i n e . each l i n e . t i o n p e r io d . The t o t a l and d a i l y d e a th r a t e d i f f e r e d fo r The s u s c e p t i b l e l i n e s were c h a r a c t e r iz e d by a sh o r t in c u b a ­ L e s s s u s c e p t i b l e l i n e s showed a more u n ifo rm d i s t r i b u t i o n o f m o r t a l i t y o v e r t h e e n t i r e p e r io d * A lth o u g h t h e nJ" and nKn g e n e r a t io n s a r e not i d e n t i c a l , t h e com p arison o f t h e p e r c e n ta g e o f s u r v iv o r s i n s im ila r l i n e s t h a t had common f a m i l i e s w it h in t h e tw o g e n e r a t io n s showed t h a t t h e r e was no s i g n i f i c a n t d i f f e r e n c e b etw een t h e tw o g e n e r a t io n s i n reg a rd t o l i n e and s e x . However, when a l l t h e b ir d s i n a g iv e n fa m ily w it h in t h e tw o g e n e r a t io n s w ere grouped t o g e t h e r and com pared t o o th e r f a m i l i e s w it h in t h e i r r e s p e c t i v e l i n e s , d i f f e r e n c e s w ere found b etw een f a m i l i e s i n some l i n e s * T h is may be e x p e c t e d a s in b r e d l i n e s have n o t as y e t rea ch ed t h e d e g r ee o f h o m o g en eity w here a l l t h e f a m i l i e s w i t h i n t h e l i n e r e a c t i d e n t i c a l l y . The m ain d i f f e r e n c e b e tw e en th e "J" and ”K” g e n e r a t io n s was e v id e n t when th e a v e r a g e le n g t h o f s u r v iv a l fo r ea ch l i n e was compared f o r t h e two g e n e r a t io n s * The ”t M t e s t s showed t h a t t h e r e 7/as a s i g n i f i c a n t - 53 - d i f f e r e n c e b etw een s e x e s when t h e "two g e n e r a t io n s w ere com pared, u s in g t h e a v e r a g e s u r v iv a l tim e a s t h e b a s is fo r com parison* T h is seem s t o be t h e m ain f a c t o r w h ich brought out t h e s i g n i f i c a n t d i f f e r e n c e b etw een t h e tw o g e n e r a t io n s when s e x e s w ere grouped to g e t h e r * v a r ia n c e ( t a b le The a n a l y s i s o f 9) f o r g e n e r a t io n WJ M showed a h i g h ly s i g n i f i c a n t d i f f e r e n c e b e tw e en t h e s e x e s and b etw een t h e l i n e s f o r a v era g e s u r v iv a l tim e • I n g e n e r a t io n nKn ( t a b le 1 0 ) t h e r e was a h ig h ly s i g n i f i c a n t d i f f e r e n c e b etw een t h e l i n e s * T here was a l s o a s i g n i f i c a n t d if f e r e n c e i n t h e i n t e r a c t i o n w h ich means s e x d i f f e r e n c e s w ere n o t th e same fo r a l l th e lin e s * The a n a l y s i s f o r g e n e r a t io n !lJ n d id n o t show a s i g n i f i c a n t d i f f e r e n c e i n t h e i n t e r a c t i o n b etw een s e x and l i n e s . A d e fin ite c o n c lu s io n c a n n o t be rea ch ed t o a cco u n t fo r t h i s s i g n i f i c a n t d if f e r e n c e i n t h e i n t e r a c t i o n i n “K” g e n e r a tio n * The r e s u l t s o f t h e d a ta c o l l e c t e d i n t h e g r o ss p a t h o lo g ic a l r e a c t io n o f t h e b r e a s t showed t h a t t h e t r a n s p la n t had grown i n t h e b r e a s t o f a l l t h e b ir d s e x c e p t t h r e e * Some o f t h e b ir d s r e c o v e r e d w h ile o th e r s d ie d a f t e r v a r i a b l e in c u b a t io n p e r i o d s . The r e a c t io n s o f t h e h o s t s b e a r in g t h e s e lym p hoid tum ors seem t o f a l l in t o a c e r t a i n p a t t e r n w h ich can be i n t e r p r e t e d a s a r e s u l t o f t h e i n t e r p l a y b etw een t h e a c t i v i t y o f t h e im p la n t and t h e r e s i s t a n c e o f t h e h o s t - T h u s, t h e r e s p e c t iv e r e s p o n s e s o f no g r o w th , d e g r e e o f g r o w th , r e g r e s s i o n , and d e g r e e o f m e t a s t a s is r e p r e s e n t e i t h e r t h e r e s i s t a n c e i n th e h o s t , a c t i v i t y o f t h e tumor t r a n s p l a n t , or c o m b in a tio n o f both* The d i f f e r e n c e i n th e p e r c e n ta g e o f m o r t a l i t y b e tw e e n b ir d s ex p o sed t o t h e n a t u r a lly o c c u r r in g d is e a s e and t h o s e g iv e n t h e tumor t r a n s p la n t p r o b a b ly i s i n th e s t a n d a r d iz a t io n o f t h e d o se in t a k e under t h e e x p e r im e n ta l c o n d it io n s w hich under n a tu r a l - 54 - c o n d it io n s i s p r a c t i c a l l y u n c o n t r o lle d - I n t h e s e e x p e r im e n ts t h e i n f e c t i v e d o se was r e l a t i v e l y s t a b l e , but t h e i n f e c t i v e u n it s c o n ta in e d i n t h e in o c u lu m we r e not known. I t t o o k t h e t r a n s p la n t some tim e t o grow t o a s i z e t h a t co u ld be d e te c te d * A p e r io d o f s i x days was not enough f o r t h e tum or t o be d e t e c t e d i n m ost o f t h e b ir d s * The t o t a l p e r c e n ta g e s o f th e b ir d s t h a t d id not show tum or growth on t h e s i x t h day a f t e r t r a n s p la n t a t io n i n g e n e r a t io n WJ W w ere 7 0 .1 , 9 2 .1 , 6 4 . 2 , 6 6 . 1 , 8 0 . 9 , 6 3 .3 and 8 0 .4 i n l i n e s 6 , 7 , 9 , 1 0 , 1 1 , 1 4 , and 15 r e s p e c t i v e l y . i n g e n e r a t io n The same p e r c e n ta g e s w ere 9 8 .5 , 8 9 . 6 , 9 0 .5 , 8 8 . 2 , 9 5 .3 , 9 0 .9 , and 9 9 .2 i n l i n e s 4 , 6 , 7 , 9 , 1 1 , 1 4 , and 15 r e s p e c t i v e l y . The t e n t h day proved t o be th e optimum day fo r tumor o b s e r v a t io n b e c a u s e more t h a n 70 p e r c e n t o f t h e b ir d s i n g e n e r a t io n s 11J" and UK” show ed tumor grow th a t t h i s t im e . The amount o f tum or grow th i n th e b r e a s t showed a w id e v a r i a t i o n b etw een d i f f e r e n t l i n e s on th e same d a y . The p e r c e n ta g e o f r e a c t o r s in ea ch c l a s s o f tu m or grow th was n ot i n d i c a t i v e o f s u s c e p t i b i l i t y or r e s i s t a n c e . A s u s c e p t i b l e and a r e s i s t a n t i n d iv id u a l d id i n some c a s e s show t h e same d e g r ee o f tumor r e a c t io n on t h e same d a y . I n g e n e r a l, t h e b ir d s t h a t l i v e d th e l o n g e s t , e i t h e r r e s i s t a n t or s u s c e p t i b l e , had t h e l a r g e s t tum ors and c o n s id e r a b le n e c r o t ic m a t e r ia l was f r e q u e n t ly p r e s e n t . For t h e mo sb p a r t when th e tum ors reach ed maximum s i z e a t abou t 17 t o 21 days t h e r e was e i t h e r a g ra d u a l r e g r e s s i o n o f t h e tu m ors or t h e b ir d s h a v in g su ch e n la r g e d tum ors d i e d . The most s u s c e p t i b l e c h ic k e n s , n o t a b ly l i n e 1 5 , w hich d ie d so o n a f t e r i n o c u l a t i o n , showed o n ly a m od erate grow th o f tu m o r and had l i t t l e m a te r ia l. In lin e or no n e c r o t i c 7 , s i m i l a r l y t o l i n e 1 5 , m o r t a lit y was v e r y h ig h e x c e p t t h a t t h e r e was a g r e a t e r v a r i a t i o n i n age a t d e a th . - 55 - I n g e n e r a l "the i n t e r n a l m e t a s t a t ic r e a c t i o n was p a r a l l e l t o t h e b r e a s t r e a c t io n * A g e n e r a l h yp erem ia o f th e i n t e r n a l organ s was p r e s e n t i n b ir d s t h a t d ie d b e fo r e t h e second w eek a f t e r in o c u la t io n * T h is was fo llo w e d by t h e fo r m a tio n o f a d e f i n i t e tumor t i s s u e * lym p h oid t i s s u e The shewed a te n d e n c y t o c o a le s c e a s th e b ir d l i v e d lo n g e r * T h is may h ave b een th e r e s u lt o f t h e n a t u r a l d evelop m en t o f tumor t i s s u e or a c r i t e r i o n o f t h e d e f e n s iv e power o f t h e b o d y . The i n t e r n a l o rg a n s t h a t showed t h e m ost m e t a s t a t ic r e a c t i o n w ere l i v e r , k id n e y , s p le e n , h e a r t , p r o v e n t r i c u l u s , p a n c r e a s , and g o n a d s. D ata on l e s i o n in c id e n c e o f t h e tum or i n o th e r organ s w ere not c o l l e c t e d , b u t t h e a d r e n a ls were in v o lv e d i n some c a s e s • T h ese e x p e r im e n ta l r e s u l t s s h o u ld be e n c o u r a g in g t o th e b r e e d e r s i n t e r e s t e d i n r e d u c in g t h e in c id e n c e o f t h i s d i s e a s e by means o f s e l e c t i o n and p o s s i b l y l i n e b r e ed in g * The f a c t t h a t t h e r e i s a p a r a l l e l i s m b etw een ft s u s c e p t i b i l i t y or r e s i s t a n c e under th e n a t u r a l d is e a s e o c cu rr e n c e and in d u c ed d i s e a s e may make t h e lo n g t a s k o f s e l e c t i o n fo r r e s i s t a n c e t o ly m p h om atosis e a s ie r * A r e a s o n a b le p la n o f s e l e c t i o n and b r e e d in g w it h in f lo c k s s u b j e c te d t o t h e d i s e a s e , may r e s u l t i n r e l a t i v e l y r e s i s t a n t f lo c k s * T h is may be th e b e s t answ er on hand t o red u ce t h e l o s s e s from t h e d is e a s e u n t i l d e f i n i t e p r o p h y la c t ic and m e d ic a l m easu res a re p r o v id e d * - 56 - SUMMARY 1* In b r e d W hite L eghorn c h ic k e n s r e s i s t a n t t o and s u s c e p t i b l e t o n a t u r a l l y o c c u r r in g lym p h om atosis ivere in o c u la t e d I n tr a m u s c u la r ly w ith m a t e r ia l from, tum or s t r a i n RPL 1 6 . 2. A t o t a l o f 2027 c h ic k e n s b e lo n g in g t o tw o d i f f e r e n t g e n e r a t io n s , "J" and "K", was u sed i n t h e s e s t u d i e s . I n g e n e r a t io n "J” random sam p les o f c h ic k e n s from in b re d l i n e s 6 , 7 , 9 , 1 0 , 1 1 , 1 4 , and 15 were u s e d . In g e n e r a t io n "K” c h ic k e n s from l i n e s 4 , 6 , 7 , 9 , 1 0 , 1 4 , and 15 were in v o lv e d . 3. S tan d ard in o cu lu m o f 0 .1 cc o f n o n - f i l t e r e d tumor c e l l s u s p e n s io n from s t r a i n RPL 16 was t r a n s p la n t e d i n t h e b r e a s t m u scle o f 2 1 -d a y o ld c h ic k e n s • 4. O b s e r v a tio n s for tumor r e a c t io n w ere made on e a ch b ir d d a i l y th r o u g h t h e 2 8 -d a y e x p e r im e n ta l p e r io d f o llo w in g i n o c u l a t i o n . 5. P e r c e n ta g e o f s u r v iv a l and a v e r a g e age a t s u r v iv a l f o llo w in g in o c u l a t i o n w it h tum or t r a n s p la n t s w ere t h e c r i t e r i a used t o m easure t h e r e s i s t a n c e and s u s c e p t i b i l i t y o f t h e c h ic k e n s i n th e v a r io u s l i n e s . 6. s e le c te d Under t h e n a t u r a l l y o c c u r r in g d is e a s e l i n e s 6 and 10 were fo r r e s i s t a n c e and l i n e s 7 , 9 , 1 1 , 1 4 , and 15 w ere s e l e c t e d fo r s u s c e p t ib ilit y . 7. Under t h e c o n d i t i o n o f t h e s e e x p e rim en ts l i n e 10 was s u s c e p t i b l e t o tum or t r a n s p l a n t s . The r e s t o f t h e l i n e s i n b o th g e n e r a t io n s w ere s i m i l a r i n t h e i r r e a c t io n t o tumor t r a n s p la n t s and n a t u r a lly o c c u r r in g ly m p h o m a to sis . 8. D ea th r a t e o f th e in o c u la t e d b ir d s was about t w ic e as h ig h a s th e d e a th r a t e o f t h e i r r e s p e c t i v e s i b s ex p o sed t o t h e n a t u r a lly o c c u r r in g d i s e a s e w ith t h e e x c e p t io n o f l i n e 6 . - 57 - 9# I n g e n e r a l "there was no s i g n i f i c a n t d i f f e r e n c e b etw een t h e tw o g e n e r a t io n s when t h e p e r c e n ta g e o f s u r v iv o r s was used as a m e a su r e . T h ere was a s i g n i f i c a n t d if f e r e n c e b etw een t h e two g e n e r a tio n s when t h e a v e ra g e s u r v i v a l tim e was u sed a s th e c r i t e r i o n . 10* a wt M t e s t U sin g l i n e s t h a t had f a m i l i e s common t o b o th g e n e r a t io n s showed t h a t t h e r e was no s i g n i f i c a n t d i f f e r e n c e b etw een t h e g e n e r a t i o n s , b etw een t h e tw o s e x e s , b etw een fe m a le s i n th e tw o g e n e r a t io n s , and b etw een m a le s i n t h e tw o g e n e r a t io n s . 11. S e x e s and g e n e r a t io n s w it h in f a m i l i e s i n l i n e s 6 , 7 , 9 , 1 1 , 1 4 , and 15 w ere com b in ed . A c h i-s q u a r e t e s t showed t h a t t h e r e w ere s i g n i f i c a n t d i f f e r e n c e s i n m o r t a l i t y b etw een f a m i l i e s i n l i n e s 6 , 9 , 1 1 , 1 4 , and 1 5 . 12. U sin g t h e a v e r a g e day o f s u r v i v a l , a wt tt t e s t showed t h a t t h e r e was a s i g n i f i c a n t d i f f e r e n c e b e tw e en s e x e s i n b o th g e n e r a t io n s . 13. The method o f e x p e c te d s u b c la s s numbers was used fo r a n a ly s is of th e d a ta . The a v e r a g e s u r v iv a l tim e i n days was u sed a s t h e c r i t e r i o n fo r c o m p a r is o n . I n g e n e r a t io n "J" t h e r e was a s i g n i f i c a n t d if f e r e n c e b etw een s e x means and l i n e m ea n s. I n g e n e r a t io n ,TK” t h e r e was a s i g n i f i c a n t d i f f e r e n c e b etw een l i n e means and i n th e s e x - l i n e i n t e r a c t i o n . 14. B r e a s t r e a c t i o n s o c cu rr e d i n 2024 b ir d s out o f a t o t a l o f 2 0 2 7 . 15. S iz e o f tumor grow th i n t h e b r e a s t or d e g r ee o f m e t a s t a s is i n ea ch o f l i v e r , k id n e y , s p l e e n , h e a r t , p r o v e n t r ic u lu s , p a n c r e a s , and g o n a d s, d id not i n d i c a t e r e s i s t a n c e or s u s o e p t i b i l i t y o f t h e in d iv id u a l b ir d • 16. The l i v e r , k id n e y , s p l e e n , h e a r t , p r o v e n t r ic u l u s , p a n c r e a s , and gonads w ere t h e organ s t h a t showed m ost o f t h e i n t e r n a l m e t a s t a s i s . - 58 - 17* The l i v e r showed tw o t y p e s o f in v o lv e m e n t, a d i f f u s e t y p e , and a c o m b in a tio n o f b o th d i f f u s e and f o c a l tum ors* The p e r c e n ta g e o f b ir d s t h a t showed t h e d i f f u s e ty p e was v e r y h ig h * 18* The s e v e r i t y o f m e t a s t a s is was i n g e n e r a l p a r a l l e l w it h b r e a s t r e a c t io n * 19* The r e s u l t s o f t h e s e e x p e r im e n ts te n d t o i n d i c a t e t h a t i t may be p o s s i b l e t o h a s t e n t h e s e l e c t i o n o f in b r e d b ir d s tow ard r e s i s t a n c e by u s in g r e s u l t s o f in o c u l a t i o n o f s ib s w ith tumor o e l l s as a c r it e r io n fo r s e l e c t i o n . - 59 - REFERENCES A c k e r b , J . E . and S . A* E d g a r, 1938* G o b le t o e l l s and age r e s i s t a n c e t o p a r a s it is m . J . P a r a s it o l. 26s 1 3 -1 4 . A sm undson, V . S . and J . B i e l y , 1 9 3 2 . I n h e r it a n c e o f r e s i s t a n c e t o p a r a l y s i s (n eu rolym p h om atosis g a llin a r u m ) . I . D if f e r e n c e s i n s u s c e p t i b i l i t y . C anadian J . R e se a r c h 6 : 1 7 1 -1 7 6 . B i e l y , J . , V . E l v ir a Palm er and V . S . Asmundson, 1 9 3 2 . I n h e r it a n c e o f r e s i s t a n c e t o fo w l p a r a ly s is (n eu ro ly m p h o m a to sis g a llin a r u m ) • II. On a s i g n i f i c a n t d i f f e r e n c e i n t h e in c id e n c e o f fo w l p a r a ly s i s i n tw o grou p s o f c h i c k s . C anadian J . R e se a rc h 6 :3 7 4 - 3 8 0 . B i e l y , J . , V . E l v i r a P a lm er, I . M. L ern er and V . S . Asmundson, 1 9 3 3 . I n h e r it a n c e o f r e s i s t a n c e t o fo w l p a r a l y s i s (n eu rolym p h om atosis g a llin a r u m ) • S c ie n c e 7 8 : 4 2 . B ittn e r , J . J . , 1938. B io l. 1 3 : 5 1 -6 4 . The g e n e t i c s o f c a n c e r i n m ic e . B ittn e r , J . J . , 1939. B r e a st c a n c e r i n m ic e . Q uar. R ev . Am. J . Cancer 3 6 : 4 4 - 5 0 . B la k em o re, F . and R . E* G lo v e r , 1 9 3 5 . Fowl p a r a l y s i s . I I . E x p erim en ts on t r a n s m i s s i o n . Cambridge U n i v ., I n s t . Anim al P a t h . R ept • D i r . 4 : 5 1 -6 4 . B u r m e ste r , B . R . and C. 0 . P r i c k e t t , 1 9 4 3 . The d evelop m en t o f h ig h ly m a lig n a n t tu m or s t r a i n s from n a t u r a ll y o c c u r r in g a v ia n lym p h om atosis • C ancer r e s e a r c h 5 : 6 5 2 -6 6 0 . B u r m e ste r , B. R . and G. E . C o t t r a l , 1 9 4 9 . U n p u b lish ed r e s u l t s . C o le , R . K ., 1 9 4 1 . G e n e tic r e s i s t a n c e t o t r a n s m i s s i b le sarcom a i n th e f 07/1 • C ancer r e s e a r c h 1 : 7 1 4 -7 2 0 . C o le , R . K ., 1 9 4 9 . The eg g and a v ia n l e u c o s i s . P o u lt r y S c i . 2 8 : 3 1 - 4 4 . C o t t r a l , G . E . , B . R . B urm ester and N e ls o n F . W a te r s, 1 9 4 9 a . The t r a n s m is s i o n o f v i s c e r a l lym p h om atosis w it h t i s s u e s from em bryonated e g g s and c h ic k s from "normal" p a r e n t s . P o u lt r y S c i . 2 8 : 7 6 1 . C o t t r a l , G. E . , 1 9 4 9 b . A v ia n ly m p h o m a to sis, a n o th e r e g g -b o r n e d i s e a s e . P r o c . 53rd A nnual M eetin g U« S . L iv e s t o c k S a n ita r y A s s o c i a t i o n , pp- 1 8 3 -1 9 2 . C o t t r a l , G. E . and B . R . B u rm ester, 1 9 4 9 c . U n p u b lish ed r e s u l t s . Crew, F . A . E . , 1 9 2 3 . The s i g n i f i c a n c e o f an a c h o n d r o p la s ia - lik e c o n d it io n met 7/ i t h i n c a t t l e . P r o c . 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